Associations of Suicide Rates and Affective Disorders with Lithium Levels in Drinking Water

LITHUANIAN UNIVERSITY OF HEALTH SCIENCES MEDICAL ACADEMY

Vilma Liaugaudaitė

ASSOCIATIONS OF SUICIDE RATES AND AFFECTIVE DISORDERS WITH LITHIUM LEVELS IN DRINKING WATER

Doctoral Dissertation Medical and Health Sciences, Public Health (M 004)

Kaunas, 2021 Dissertation has been prepared at the Neuroscience Institute (until 2017.12.31 Behavioral Medicine Institute) of Medical Academy of Lithuanian University of Health Sciences during the period 2016–2021.

Scientific Supervisor Prof. Dr. Nida Žemaitienė (Lithuanian University of Health Sciences, Medical Academy, Medical and Health Sciences, Public Health – M 004).

Consultant Prof. Dr. Narseta Mickuvienė (Lithuanian University of Health Sciences, Medical Academy, Medical and Health Sciences, Nursing – M 005).

Dissertation is defended at the Public Health Research Council of the Lithuanian University of Health Sciences:

Chairperson Prof. Dr. Ričardas Radišauskas (Lithuanian University of Health Sciences, Medical Academy, Medical and Health Sciences, Public Health – M 004).

Members: Prof. Dr. Žemyna Milašauskienė (Lithuanian University of Health Sciences, Medical Academy, Medical and Health Sciences, Public Health – M 004); Assoc. Prof. Dr. Rugilė Ivanauskienė (Lithuanian University of Health Sciences, Medical Academy, Medical and Health Sciences, Public Health – M 004); Prof. Dr. Valentina Vengelienė (Vilnius University, Natural Sciences, Biology – N 010); Prof. Dr. Gil Zalsman (Tel Aviv University, Medical and Health Scien- ces, Medicine – M 001).

Dissertation will be defended at the open session of the Public Health Research Council on the 9th of April, 2021 at 12 a. m. in the auditorium A-202 of the Centre for the Advanced Pharmaceutical and Health Technologies of Lithuanian University of Health Sciences. Address: Sukilėlių 13, LT-50166 Kaunas, Lithuania. LIETUVOS SVEIKATOS MOKSLŲ UNIVERSITETAS MEDICINOS AKADEMIJA

Vilma Liaugaudaitė

SAVIŽUDYBIŲ IR AFEKTINIŲ SUTRIKIMŲ SĄSAJOS SU LIČIO KONCENTRACIJA GERIAMAJAME VANDENYJE

Daktaro disertacija Medicinos ir sveikatos mokslai, visuomenės sveikata (M 004)

Kaunas, 2021 Disertacija rengta 2016–2021 m. Lietuvos sveikatos mokslų universiteto Medicinos akademijos Neuromokslų institute (iki 2017-12-31 Elgesio medicinos institutas).

Mokslinė vadovė prof. dr. Nida Žemaitienė (Lietuvos sveikatos mokslų universitetas, Medicinos akademija, medicinos ir sveikatos mokslai, visuomenės sveikata – M 004).

Konsultantė prof. dr. Narseta Mickuvienė (Lietuvos sveikatos mokslų universitetas, Medicinos akademija, medicinos ir sveikatos mokslai, slauga – M 005).

Disertacija ginama Lietuvos sveikatos mokslų universiteto Visuomenės sveikatos mokslo krypties taryboje:

Pirmininkas prof. dr. Ričardas Radišauskas (Lietuvos sveikatos mokslų universitetas, Medicinos akademija, medicinos ir sveikatos mokslai, visuomenės sveikata – M 004).

Nariai: prof. dr. Žemyna Milašauskienė (Lietuvos sveikatos mokslų universitetas, Medicinos akademija, medicinos ir sveikatos mokslai, visuomenės sveikata – M 004); doc. dr. Rugilė Ivanauskienė (Lietuvos sveikatos mokslų universitetas, Medicinos akademija, medicinos ir sveikatos mokslai, visuomenės sveikata – M 004); prof. dr. Valentina Vengelienė (Vilniaus universitetas, gamtos mokslai, biologija – N 010); prof. dr. Gil Zalsman (Tel Avivo universitetas, medicinos ir sveikatos mokslai, medicina – M 001).

Disertacija ginama viešame Visuomenės sveikatos mokslo krypties tary- bos posėdyje 2021 m. balandžio 9 d. 12 val. Lietuvos sveikatos mokslų universiteto Naujausių farmacijos ir sveikatos technologijų centro A-202 audi- torijoje. Adresas: Sukilėlių pr. 13, LT-50166 Kaunas, Lietuva. CONTENT

LIST OF ABBREVIATIONS ...... 7 DEFINITIONS OF TERMS ...... 8 INTRODUCTION ...... 10 AIM AND OBJECTIVES ...... 12 1. REVIEW OF LITERATURE ...... 14 1.1. Epidemiology of suicide ...... 14 1.1.1. Suicide Worldwide ...... 14 1.1.2. Suicide in Lithuania ...... 15 1.1.3. Suicide prevention ...... 15 1.1.4. Suicide and mental and behavioral disorders ...... 17 1.2. The Anti-suicidal effect of lithium ...... 19 1.2.1. Dietary source of lithium ...... 19 1.2.2. The Effects of lithium deficiency on behavioral parameters ...... 20 1.2.3. Mechanistic considerations of lithium ...... 20 1.3. Summary of studies investigating the anti-suicidal effects of lithium in drinking water ...... 21 1.3.1. Analysis of selected studies ...... 21 1.3.2. Studies on potential anti-suicidal effects of lithium as a trace element in drinking water ...... 26 1.3.3. Summary of the literature review ...... 27 2. MATERIAL AND METHODS ...... 29 2.1. Study design ...... 29 2.2. Methods ...... 30 2.2.1. Study area: evaluation of lithium levels in drinking water ...... 30 2.2.2. Dependent variable: suicide standardized mortality rate ...... 31 2.2.3. Independent variables ...... 31 2.3. Statistical analysis ...... 32 3. RESULTS ...... 36 3.1. Lithium levels in drinking water, suicide SMRs, and socio-demographic characteristics in the municipalities ...... 36 3.2. Association of suicide SMR with lithium levels in drinking water: Study I (Pilot study) ...... 39 3.3. Association of suicide SMR with lithium levels in drinking water: Study II ...... 42 3.3.1. The correlates of suicide SMRs ...... 42 3.3.2. Multiple regression analysis of suicide SMRs ...... 43

5 3.4. Relationships between suicide SMRs, incidence of affective disorders, and lithium levels ...... 48 3.4.1. Analysis of suicide SMRs and other indicators of municipalities according to quartiles of lithium levels...... 48 3.4.2. Analysis of suicide SMRs and other indicators of municipalities according to a median of lithium levels ...... 49 3.4.3. Associations of suicide SMRs, and mental and behavioral disorders with lithium levels in low lithium exposure group ...... 53 3.4.4. Relationships between suicide SMRs, mental and behavioral disorders, and lithium levels in high lithium exposure group ...... 55 3.4.5. Exploratory factor analysis of the variables in high lithium exposure group: supplementary analysis ...... 60 4. DISCUSSION ...... 63 4.1. The lithium level in drinking water of the central wellfields in Lithuanian municipalities ...... 63 4.2. The association between suicide SMR and lithium levels in drinking water and across municipalities ...... 64 4.2.1. Association between suicide SMR and lithium levels in drinking water: Study I (Pilot study) ...... 64 4.2.2. Association between suicide SMR and lithium levels in drinking water: Study II ...... 65 4.3. The impact of incidence of affective disorders on an association of suicide SMR with lithium level in drinking water ...... 68 LIMITATIONS AND STRENGTH OF THE STUDY ...... 75 CONCLUSIONS ...... 77 PRACTICAL AND SCIENTIFIC RECOMMENDATIONS ...... 78 SANTRAUKA ...... 80 REFERENCES ...... 106 PUBLICATIONS ON THE DISSERTATION THEME ...... 117 ANNEXES ...... 137 Annexe 1 ...... 138 Annexe 2 ...... 139 Annexe 3 ...... 140 CURRICULUM VITAE ...... 141 ACKNOWLEDGEMENTS ...... 143

6 LIST OF ABBREVIATIONS

AUD – Alcohol use disorder BD – Bipolar disorder CI – Confidential interval DALYs – Disability Adjusted Life Years DDD – Defined Daily Dose EU – European Union IQR – Interquartile range Li – Lithium MBD – Mental and behavioral disorders MDD – Major depressive disorder OECD – Organization for Economic Cooperation and Development OR – Odds ratio PC – Principal components PCA – Principal component analysis RDA – Recommended Dietary Allowances RR – Relative risk SA – Suicidal attempt SB – Suicidal behavior SD – Standard deviation SGA – Second generation antipsychotics SI – Suicidal ideation SMR – Standardized mortality rates SPSS – Statistical Package for the Social Sciences USEPA – United States Environmental Protection Agency VIF – Variance Inflation Factor WHO – World Health Organization WLS – Weighted Least-Squares

7 DEFINITIONS OF TERMS

Affective disorder – any mental disorder, as depressive disorder, bipolar disorder in which a major disturbance of feelings or emotions is predominant. Anxiety disorder – chronic feelings of overwhelming anxiety and fear, unattached to any obvious source, that can grow progressively worse if not treated. The anxiety is often accompanied by physical symptoms such as sweating, cardiac disturbances, diarrhea or dizziness. Bipolar disorder – a serious illness that causes shifts in a person’s mood, energy and ability to function. Dramatic mood swings can move from “high” feelings of extreme euphoria or irritability to depression, sometimes with periods of normal moods in between. Comorbidity – in general, the existence of two or more illnesses, whether physical or mental, at the same time in a single individual. Depression – mental state characterized by feelings of sadness, lone- liness, despair, low self-esteem, and self-reproach; accompanying signs include psychomotor retardation or, at times, agitation, withdrawal from interpersonal contact, and vegetative symptoms, such as insomnia and anorexia. The term refers to a mood that is so characterized or to a mood disorder. Mental disorder – psychiatric illness or disease whose manifestations are primarily characterized by behavioural or psychological impairment or function, measured in terms of deviation from some normative concept; associated with distress or disease, not just an expected response to a particular event or limited to relations between a person and society. Lithium – lithium carbonate or other preparations of lithium metal used to treat manic depression and bipolar disorders. Psychosis – mental disorder in which the thoughts, affective response, ability to recognize reality, and ability to communicate and relate to others are sufficiently impaired to interfere grossly with the capacity to deal with reality; the classic characteristics of psychosis are impaired reality testing, hallucinations, delusions, and illusions. Schizophrenia – a mental illness that is characterized by disturbances in thought (such as delusions), perception (such as hallucinations), and behavior (such as disorganized speech or catatonic behavior), by a loss of emotional responsiveness and extreme apathy, and by noticeable deterioration in the level of functioning in everyday life. Substance use disorder – any mental or behavioral disorder resulting from the use of one or more psychoactive substances, whether or not medically prescribed. Suicidal ideation – thoughts or act of taking one’s own life.

8 Suicide – a self-inflicted death with evidence (either explicit or implicit) of intent to die. Suicide attempt – is defined as a self-inflicted, potentially injurious behavior with a nonfatal outcome for which there is evidence (either explicit or implicit) of intent to die. A suicide attempt may result in no injuries, injuries, or death.

9 INTRODUCTION

Suicide is a serious global public health problem that demands our attention but preventing suicide is no easy task [1]. Suicide is the second leading cause of death among 15–29 year olds globally and one of the most important indicators of the public’s state of mental health [2]. Suicide is a complex phenomenon with many contributing factors including psychological, social, economic, biological, and environmental [3– 6]. However, mental health disorders are also attributed to a significant number of indirect deaths through suicide and self-harm. Suicide deaths are strongly linked, although not always attributed to mental health disorders. There is strong evidence that suicide mostly occurs among people with affective disorders [7]. The suicide risk has been estimated at 6–10% in the affective disorder population, which is 10 times the corresponding risk in non- psychiatric populations [8]. Treatment of patients with suicidal behavior is one of the most challenging tasks for health-care professionals [9]. To date, several strategies have been proposed for suicide prevention, both at population and individual level, some of which may be pharmacological [10, 11]. Lithium is the “gold standard” mood stabilizer against which potential mood stabilizer agents are judged [12]. A meta-analysis demonstrated an overall significant efficacy of lithium in preventing suicide with a highly significant reduction in rates of suicide [13–16]. In clinical practice it is well established that lithium has a mood-stabilizing and suicide-preventive effect in individuals suffering from affective disorders [8, 15]. A systematic review and meta-analysis of randomized trials on this topic suggested that lithium treatment reduces mortality and suicide by more than 60% in people with major depression or bipolar disorder [17]. A placebo-controlled trial data showed that low doses of lithium might improve and stabilize mood quite rapidly in former drug users [18]. The duration of lithium pharmacotherapy needed for suicide prevention is important. Findings from the international multi-centre trial are compatible with the idea that long-term lithium treatment extends the survival of patients suffering from affective disorders to match the general population. The randomized as well as open clinical trials revealed that the risks of completed and attempted suicide were consistently lower, by approximately 80%, during treatment of bipolar and other major affective disorder patients with lithium for an average of 18 months [19]. Although trace lithium intake doses are significantly lower than those used for the treatment of patients with psychiatric disorders [20, 21], there is

10 growing evidence that even very low lithium levels induced by routine consumption of lithium from tap water may have anti-suicidal effects both in patients with affective disorders, and in the general population [22, 23]. One hypothesis the explaining anti-suicidal effects of low lithium levels is that long-term exposure to lithium through routinely drinking water may mitigate low absolute lithium levels [24]. In addition, lithium appears to have value in augmenting antidepressant treatment. Lithium continues as the standard and most extensively evaluated treatment for bipolar disorder, especially for long- term prophylaxis [8, 14, 19]. Several epidemiologic studies have reported that lithium in drinking water may be associated with lower rates of suicide mortality [24–31], lower incidence of dementia, lower levels of adolescents' depression, aggression [32, 33] and psychotic experiences [34] at the population level. The biological actions of lithium are complex and not fully understood. Lithium has been the subject of suicide research for a long time [7, 35, 36]. In addition to its mood-stabilizing, anti-depressive and anti-manic effects in individuals with bipolar and other major affective disorders, some studies suggest that lithium in therapeutic doses has an anti-suicidal effect [8, 14, 37– 39]. This may be mediated through its mood-stabilizing properties or through a reduction of aggressiveness and impulsivity, which are associated with increased risk of suicide [33]. Despite this interest, none of the studies, to the best of our knowledge has studied the interaction between the suicide rates, prevalence of mental and behavioral disorders and naturally occurring lithium in drinking water. Based on the existing knowledge we hypothesized that higher levels of lithium in drinking water may exert an anti-suicidal effect in the population with high incidence of affective disorders.

11 AIM AND OBJECTIVES

The aim of this study was to establish whether suicide rates and incidence of affective disorders are associated with lithium levels in drinking water. Objectives of the study: 1. To examine the lithium levels in drinking water in Lithuanian municipal central wellfields. 2. To investigate the association of suicide mortality rates with lithium levels in drinking water. 3. To evaluate whether incidence of affective disorders is associated with lithium levels in drinking water. Statements of the hypotheses We hypothesized, that: 1. Lithium levels in drinking water differs across the municipalities; 2. Increased lithium levels in drinking water may be associated with a lower suicide mortality rate; 3. Lithium levels in drinking water are negatively associated with suicide rates in the population with higher incidence of affective disorders. Scientific novelty of the study Firstly, this is the first ecological study on the association between lithium levels in the public drinking water and suicide rates in Eastern Europe. This study investigated the lithium level in drinking water wellfields across the Lithuanian municipalities, and tested the hypothesis that higher lithium levels in the public drinking water are associated with lower suicide mortality rates. Secondly, we evaluated the association of mental and behavioral disorders with lithium level in drinking water. Our study revealed a higher incidence of affective disorder in municipalities with higher lithium level in drinking water. Therefore, it seems probable that the anti-suicidal effect of lithium may be unrelated to the mood-stabilising effects and that very low lithium levels (comparing with therapeutic doses) may possess an anti- suicidal effect. On the other hand, although lithium levels in drinking water are extremely low, long-term exposure to lithium may be a factor which mitigates low absolute levels. The study results demonstrate a new knowledge in understanding the association of suicide rates with lithium levels in drinking water linked to incidence of affective disorders.

12 Finally, our study might provide new knowledge for future research studies evaluating biological mechanisms on suicidal behavior at the population level, especially in patients with affective disorders. Contribution of the author The author of the dissertation has been actively involved in the collection of water samples since the beginning of the study. She has been in direct contact with the heads of municipalities’ wells regarding the permission to collect water samples for lithium testing. In the course of the research, the author prepared a project to obtain funding for research of the central wells of Lithuanian municipalities. She also cooperated with the representatives of the Institute of Geology and Geography of Nature Research Center for the collection of water samples, and the representatives of the laboratory for the analysis of drinking water samples. The author collected and processed socio-demographic, incidence and mortality indicators of Lithuanian municipalities, using the database of direct and indirect access to Lithuanian health indicators. She processed the results and performed statistical analysis and interpretation of the data. During the doctoral studies, the author presented the results of the researches at international and national conferences in Lithuania and abroad. The obtained results were published in peer-reviewed foreign journals.

13 1. REVIEW OF LITERATURE

1.1. Epidemiology of suicide

1.1.1. Suicide Worldwide Globally, suicides are the second leading cause of premature mortality in individuals aged 15 to 29 years (preceded by traffic accidents), and number three in the age-group 15–44 years [40–42]. According to recent review by Bachmann et al (2018) crude and age-standardized suicide rates according to WHO regions, amount to 10.7 per 100,000 worldwide but varies in certain regions [43]. The Eastern Mediterranean region has suicide rates of 3.8 and 4.3, the African region 8.8 and 12.8, the Americas 9.6 and 9.1, the Western Pacific region 10.8 and 9.1, South East Asia 12.9 and 13.3, and Europe 14.1 and 11.9 (all crude and age-standardized) [44]. The highest suicide rates for both men and women are found in Europe, more particularly in Eastern Europe, in a group of countries that share similar historical and sociocultural characteristics, such as Estonia, Latvia, Lithuania and, to a lesser extent, Finland, Hungary and the Russian Federation [40]. Obviously, the European area presents the highest absolute or crude suicide rate, namely above the global suicide rate of 10.7 for both genders [44, 45]. This is the case despite the fact that since 1980, a drop in suicide rates was reached through preventive measures and assisted suicides were taken out of the statistics. On the other hand, however, data quality is much better in comparison to other regions of the world [46]. Although there has been some progress in reducing historically high mortality rates from suicide, it nevertheless remains an important cause of death, particularly among men [47]. It was estimated that nearly three times as many men as women die by suicide in high-income countries, in contrast to low- and middle-income countries, where the rate is more equal [48, 49]. It is very difficult to obtain reliable data on suicide rates. For example, assuring rates of suicide attempts is nearly impossible, not least because a suicide attempt may not come to anyone’s attention, much less to the attention of the health care system. Nevertheless, the registration of suicides and suicide attempts is a desirable goal towards better prevention, detection, and intervention [40].

14 1.1.2. Suicide in Lithuania Although the suicide rate in Lithuania has been decreasing every year since 2000 [50], Lithuania is still of the top countries in terms of suicide rate. Lithuania ranks fourth in the world according to suicide rate, and it ranks second for the male suicide rate among all countries in the world [42, 44]. According to data provided by the Institute of Hygiene in Lithuania, 683 people committed suicide in 2018, down from 748 in 2017, 823 in 2016, 891 in 2015, 935 in 2014 and 1,085 in 2013 [50]. According to the Eurostat data, the average rate of death by suicide in the EU in 2016 was 12.85 (per 100,000 residents), while the SMR in Lithuania was 25.7 (per 100,000 residents) exceeding the European Union average two times [51, 52]. In 2018 it was at 24.4, which means it remained slowly decreased for two years [50]. The most vulnerable groups include mid-aged (45–59 years) and over 75 years’ old males living in rural area. Men are five times more likely than women to commit suicide, so municipalities are developing rapid intervention algorithms to address this trend [50, 52]. The Organization for Economic Cooperation and Development (OECD) notes that Lithuania’s high suicide rate is tied to many different factors, including rapid social and economic change which increase both psychological and social insecurity, as well as the lack of a national suicide prevention strategy [52].

1.1.3. Suicide prevention Countries and communities may influence suicide rates by measures of primary and secondary prevention [1]. In recent years, the authorities have launched a number of suicide prevention campaigns. Social and economic conditions have also changed, leading to a 45% decrease in the number of deaths due to suicide between 2000 and 2016, but it is still more than double the OECD average for the general population and nearly three-times the OECD average for men [50, 52]. One of the goals of the Lithuanian Health Strategy 2014–2025 [53] is to decrease the SMR due to suicide down to 19.5 cases per 100,000 residents by 2020, and then to 12 cases by 2025. The rates had only been reduced by 1.6% from 2012 to 2015 [54]. The achievement of such an ambitious goal necessitates mobilization and purposeful action on the part of all sectors (health, education, social security and labour, interior affairs, non-government organizations (NGO), etc.). Important efforts have been made in recent years to improve mental health services, and decrease alcohol consumption, which have contributed to a reduction in the number of deaths by suicide. Between 2016 and 2018, the government adopted a number of policies to tackle excessive alcohol

15 consumption. Measures included an increase in excise taxes, a prohibition of alcohol sales in some places such as petrol stations, an increase in the legal age for purchasing and consuming alcohol to 20 years, and a limitation of operating hours of sale [52]. Many strategies have been developed, not only to prevent suicide but also to detect depression symptoms earlier and provide more appropriate treatment for other mental health issues. Psychological help for those who are not willing or not able to seek out the professional assistance of a psychologist or a psychotherapist due to financial restrictions is available through online or phone suicide prevention hotlines. In 2017, the National Audit Office renewed efforts to identify and support individuals at risk of mental health issues and to ensure immediate and continuous support to people who had attempted suicide, emphasizing the need for information sharing between institutions [52]. On 1st November 2018, the Lithuanian Health Minister published a paper stating the procedure for providing help to suicide survivors. The document includes prepared goals to increase training of key gatekeepers, volunteers, and professionals regarding recognition of risk factors, warning signs and at- risk behaviors; the provision of effective interventions; and the development and promotion of effective clinical and professional practice to support clients, families and communities. Psychosocial assessment can be a vital tool for self-harm management, engaging patients in treatment and improving their rates of aftercare. A recent Lithuanian Health Minister order states that, from November 2019, new health care services will be provided in mental health care institutions to children and adolescents. In addition, the network of institutions providing psychiatric day care treatment will expand from five institutions to ten. Since 2019, the Ministry also started financing municipal Public Health Bureaus to promote mental health prevention in schools. The objectives of this program are to enhance the competences of school staff in detecting and addressing mental health issues and improving overall mental health literacy. According to data provided by the State Patients’ Fund, the number of visits to medical psychologists in clinics has tripled over the past three years – from 20,959 in 2014 to 65,000 in 2017. The support from other people during a suicidal crisis is one of the strongest suicide deterrents [55]. Unfortunately, individuals with suicide behavior often avoid seeking help from others [56]. There is also a problem with the reliability of suicide confirmation and reports that needs to be improved. Suicides are most commonly found misclassified according to the codes of the 10th edition of the International Classification of Diseases and Related Health Conditions (ICD-10) [49, 57]. In addition, stigma associated with suicide (and suicide attempts) has

16 profound psychological and behavioral effects on individuals and families, who may avoid seeking help, and may conceal or deny suicide. This can cause false death registration and coding practice [58, 59]. Providing case registration information to policy makers, researchers and health professionals allows greater exposure of the problem of suicide and can be a way of raising awareness, initiating research and development in prevention campaigns, and monitoring the effectiveness of suicide prevention and intervention strategies. Until now, the reasons for the marked heterogeneity in international suicide rates are not fully understood. One important observation is that national suicide rates by particular lethal methods (such as hanging, poiso- ning, gassing, shooting, jumping, and drowning) vary greatly between nations [60], but tend to be stable within nations on a year-to-year basis [61]. This predictability of method specific suicide rates underpins most universal measures to prevent suicide. Some potential universal prevention strategies come at a greater cost. Examples include better access to health care [62] and measures to reduce unemployment [63]. Other universal, potentially important measures might be inexpensive but hard to achieve, for example reducing suicide by reducing the stigma associated with accessing mental health care [64, 65]. Despite the challenges faced by Lithuania and other countries with a high suicide rate, universal measures hold the best hope for global suicide prevention.

1.1.4. Suicide and mental and behavioral disorders Suicide is not a mental health problem itself, but is linked with mental distress. The etiology of suicidal behavior and suicidal ideation is multifactorial, although one of the most common risk factors is having a psychiatric disease. Several psychological autopsy studies have supported high rates of psychiatric disorders among individuals who die by suicide [66]. Further, a meta-analysis of 3,275 suicides reported that 87.3% of suicide completers had been diagnosed with a psychiatric disorder prior to the suicide [67]. Other studies suggest that mental illness accounts for a large majority of suicides; numbers are at least 10 times as high as in the general population. Several studies from developing and industrialized countries indicate a prevalence of mental disorders in about 90% of cases of suicide; the reported percentage of completed suicides in this context ranges between 60% and 98% of all suicides [68–71]. Affective disorders are a set of psychiatric disorders, also called mood disorders. The main types of affective disorders are depression and bipolar disorder (BD). Symptoms vary by individual and can range from mild to

17 severe. One important feature of affective disorders is that they are associated with an extremely high risk of suicide, as the majority of those committing suicides are suffering from some type of affective disorder [72]. The suicide-related mortality among patients with affective disorders is approximately 30 times higher and overall mortality 2 to 3 times higher than suicide-related mortality in the general population [73]. In their meta-study of the mental health-suicide relationship, Ferrari et al. (2014) assess the pooled relative risk of suicide across a range of mental health and substance use disorders [70]. It has been reported that on average, 43.2% (SD 18.5%) of suicide cases were diagnosed with any affective disorders (including depressive and bipolar disorders) and 25.7% (SD 14.8%) with other substance problems [74]. Depression is the leading cause of death by suicide worldwide and the second cause of Disability Adjusted Life Years (DALYs) in the age category 15–44 years [70, 75]. Bradvik’s study (2018) showed that the majority of suicides are related to a psychiatric condition, including depression, substance use disorders, and psychosis [76]. Other research suggests that the suicide risk for mental disorders including depression, alcoholism, and schizophrenia is around 5% to 8%, while up to 60% of people who commit suicide have depression [77]. An individual with depression, for example, is 20 times more likely to die from suicide than someone without; someone with anxiety disorder around 3 times; schizophrenia around 13 times; bipolar disorder 6 times; and anorexia 8 times as likely. Although the total prevalence of mental health and substance use disorders does not show a direct relationship to suicide, there are notable links between specific types of mental health disorders and suicide. Longitudinal studies have consistently demonstrated the importance of past suicide attempts, cigarette smoking, alcohol use disorders, and comorbid personality disorder as predictors of future suicide attempts in depressed populations [78, 79]. The important risk factor for lifetime suicide attempt is depression, which implies that the lifetime prevalence of suicide attempts could be reduced significantly by preventing depression and by recognizing and adequately treating it [1, 80]. Advances in understanding suicidal thinking, strongly intertwined with mood states, as well as psychological and neurocognitive factors influencing a person’s decision making [81, 82] are needed to advance the clinical care of suicidal patients and prevention of suicidal behavior. The risk of suicide has previously been associated with psychiatric disorders, but new study shows that neurological disorders are also linked to an elevated suicide rate [83].

18 Among the many risk factors for suicide are mental illness, physical illness, previous suicide attempt, substance abuse, family history of suicide, impulsiveness, hopelessness, isolation, loss, relationship, social, work, and financial [84–87]. Taking into account results from previous studies, it was hypothesized that the most people who encounter such challenges learn to cope with them or find ways to overcome them, going on to survive and sometimes flourish. However, some people with limited psychological reserves who face the same challenges might come to feel that suicide, however undesirable, is preferable to living [88, 89]. Although there is no simple explanation for such counter-intuitive human behavior, social and cultural factors, media exposure, and availability of lethal means are woven in a complex web with other risk factors that can lead to suicide. The complexity of risk factors for suicide suggests that many approaches to suicide prevention should be considered and customized to accommodate local circumstances.

1.2. The Anti-suicidal effect of lithium

1.2.1. Dietary source of lithium Lithium is a trace element widely distributed on Earth [90, 91]. Because of its similarity to sodium and potassium, lithium easily crosses all biological barriers, meaning that it shows almost complete oral absorption and a uniform distribution in body fluids [91]. A study by Schrauzer reported that dietary lithium which has received scant attention is found in grains and vegetables and to some extent animal derived foods [92]. In some areas, drinking water also provides significant amounts of the element. Water moving through the ground do react to varying degrees with the surrounding minerals (and other components), and it is these rock-water interactions that give the water its characteristic chemistry. Lithium is detectable at variable concentrations in drinking water throughout the world [8, 36, 93, 94]. Human dietary lithium intakes depend on location and on the type of foods consumed, and vary over a wide range. Mobilized by weathering processes, lithium is transported into soils, from which it is taken up by plants and enters the food chain. Lithium was detected in human organs and fetal tissues in the late 19th century, leading to early suggestions of possible specific functions in the organism [90, 95]. However, it took another century before evidence for the essentiality of lithium became available. Research showed that lithium as a substance occurring naturally in food and drinking water may exert positive effects on mental health [90].

19 In studies conducted between the 1970s to the 1990s, rats and goats maintained on low lithium rations were shown to exhibit higher mortalities as well as reproductive and behavioral abnormalities. In human defined lithium deficiency, diseases have not been characterized, but low lithium intakes from water supplies were associated with increased rates of suicides, homicides and the arrest rates for drug use and other crimes [95]. Although lithium does not have a known biological role and does not appear to be an essential element for humans, the available experimental evidence now appears to be sufficient to accept lithium as essential; a provisional RDA for a 70 kg adult of 1,000 µg/day is suggested [92].

1.2.2. The Effects of lithium deficiency on behavioral parameters As lithium deficiency in humans is unlikely, any symptoms of deficiency, if at all observable, would be expected to be mild and manifest themselves primarily by behavioral rather than physiological abnormalities. Evidence linking low lithium intakes with altered behavior and aggressiveness in humans was reported by Dawson et al. [96]. These authors compared the regional mental hospital admission and homicide rates for 1967–1969 with the lithium concentrations in tap water and urine samples obtained from 24 county sites in Texas. The highest significant inverse associations of water lithium levels were observed with first mental hospital admissions for psychosis, neurosis and personality disorders. The decreasing order of magnitude of the associations was neurosis, schizophrenia psychosis, first admission, all admissions, personality, homicide and secondary admissions. Lithium deficiency may not only be caused by low dietary lithium intakes but can also be secondary to certain diseases. Research shows that organ lithium contents of kidney disease and, especially, of dialysis patients are approaching deficiency levels [14].

1.2.3. Mechanistic considerations of lithium Lithium acts on mood and suicidality via complex interactions with the serotoninergic system [67] to the decrease cerebral level of tryptophan and serotonin [66]. The research shows the important role testosterone plays in the regulation of mood and behavior, therefore it is a potentially important marker for suicide risk in an already at-risk population [67]. The biochemical mechanisms of action of lithium appear to be extra- ordinarily complex, multifactorial and strongly inter-correlated with the functions of other elements, drugs, enzymes, hormones, vitamins, growth and transforming factors. Although these were mostly observed at pharmacological levels, they could also occur at nutritional levels, accounting for the

20 unusually broad activity spectrum of lithium [11]. However, lithium has been shown to enhance folate and B12 transport into cells [67]. The transport of these factors is inhibited in lithium deficiency and can be restored by lithium supplementation. Since vitamin B12 and folate also affect mood associated parameters, the stimulation of the transport of these vitamins into brain cells by lithium may be cited as yet another mechanism of the anti-depressive, mood elevating and anti-aggressive actions of lithium at nutritional dosage levels. Recognition of the inter-correlated nature of all biological actions of lithium may result in improved therapeutic concepts. Thus, the joint admi- nistration of lithium with vitamin B12 and folate may prove more effective than lithium or the vitamins alone [14]. The fact that embryonal lithium concentrations are the highest during early fetal development suggests that it is specifically needed [96].

1.3. Summary of studies investigating the anti-suicidal effects of lithium in drinking water

Research has shown that lithium has preventive capabilities against suicide and even on a trace level, it can be therapeutic on other mental health issues. Multiple studies have attempted to find an inverse association between suicide and high trace-levels of lithium in drinking water. The electronic search strategy on the PubMed database was conducted from the date of inception of the studies in the databases to October 2019. The following keyword combinations were used: “suicide” AND “lithium in drinking water”, or “lithium in public water”. Using the keyword combinations described above, the search in PubMed provided 42 results. Studies were included if they published in English, and investigating the relationship between the level of lithium in drinking water and suicides rates in the general population. After removal of 16 duplicates, of 10 reviews, of 2 opinions of the remaining 14 records were screened for eligibility.

1.3.1. Analysis of selected studies The selected 14 papers reported studies published between 1990 and 2019 in different countries. The studies available to date on the relationship between the level of lithium in drinking water and suicide and the main results obtained are presented in Table 1.3.1.1. Although results are surprisingly concordant, methodological limitations may decrease the validity of the findings. The first reported negative association between lithium levels in tap water and suicide rates in Texas was published by Schrauzer and Shrestha

21 [95]. Using data from 27 Texas counties for the period 1978–1987, it was found that the incidence rates of suicide, homicide, and rape were significantly higher in counties whose drinking water lithium levels ranged from 70 to 160 µg/L. These results were replicated in two studies by Ohgami et al. [24] and by Kapusta et al. [26], one from Japan and one from Austria concluded that areas with higher lithium levels in the drinking water had lower suicide rates. They found that lithium levels were significantly and negatively associated with suicide standardized mortality ratio averages for the period 2002–2006 and suggested that even very low levels of lithium in drinking water may play a role in reducing suicide risk within the general population. Knudsen et al. (2017) reported that lithium levels in the drinking water of 275 municipalities in Denmark ranged from 0.6 to 30.7 μg/L, and surprisingly that there was an increasing suicide rate by increasing lithium levels [97]. These two studies had similar locations and lithium ranges but opposite findings. However, it is notable that the significant relationship between lithium levels of drinking water and the suicide rate has been proven after adjustment for some confounders. Kapusta et al. [26] evaluated the association between local lithium levels in drinking water and suicide mortality at district level in Austria. The overall suicide rate as well as the suicide mortality ratio were inversely associated with lithium levels in drinking water and remained significant after sensitivity analyses and adjustment for socioeconomic factors. However, Kabacs et al. [93], did not prove this association between lithium in drinking water and suicide rates across the East of England. The analyses of the data showed that there was no correlation between lithium levels in drinking water and suicide mortality rates in the 47 subdivisions of the East of England. This negative result could in part be explained by methodological weaknesses of the study, for example, sociodemographic or socioeconomic characteristics were not included.

22 Table 1.3.1.1. Summary of studies investigating the anti-suicidal effects of lithium in drinking water Lithium levels Number of samples References Adjusted for in drinking Results (no. of areas) water, µg/L Schrauzer and 27 counties Population density 0–160 Low lithium intakes from water Shrestha supplies were associated with 1990/Texas increased rates of suicides Ohgami et al. 18 municipalities of Population size 0.7–59 Even very low levels of lithium in 2009/Japan Oita Prefecture drinking water may play a role in reducing suicide risk within the general population Kabacs et al. 47 samples from No <1–21 No association between lithium 2011/UK East 47 subdivisions levels in drinking (tap) water and 2 3 of England mortality from suicide Kapusta et al. 99 districts Population density, per capita income, 11.3 Higher natural lithium 2011/Austria proportion of Roman Catholics, mental (SD ± 27) concentrations in drinking water are health service providers associated with lower suicide mortality rates Giotakos et al. 149 water samples Population density 11.10 Tendency for lower suicide rates in 2013/Greece from 34 prefectures (SD ± 21.16; the prefectures with high levels of 0.1–121) lithium in drinking water Blüml et al. 3123 lithium Population density, age, gender, race/ 2.8–219 Higher lithium levels in the public 2013/Texas water samples, ethnicity, median income per household, drinking water were associated with 226 counties poverty and unemployment rates lower suicide rates Sugavara et al, 40 municipalities, The density of medical institutions per 0–12.9 Significance found for females 2013 Aomori prefecture, 10,000 people and the unemployment rate before adjustments with a trend after Japan (1,373,339) adjustments (p = 0.10) and not whatsoever for males Table 1.3.1.1. Continued Lithium levels Number of samples References Adjusted for in drinking Results (no. of areas) water, µg/L Helbich et al. 6,460 lithium Population density, per capita income, 11.3 Suicide rate, SMR inversely 2015/Austria measures of proportion of Roman Catholics, density of (SD ± 27) associated with Li levels even after 99 districts psychiatrists per 10,000 population, the adjustment of lithium prescriptions number of general practitioners, the density of psychotherapists per 10,000 people Pompili et al. 145 samples Population size, mountainous area, highly 5.28 Lithium concentrations and local 2015/Italy urbanized, geographic location (0.11–60.8) suicide rates were not significantly inversely related Ishii et al, 2015 274 municipalities, Proportions: of elderly people, of 1-person 0–130 Significantly inverse associations for 2 4 Kyushu Island, households, of people with college educa- the overall SMRs (β = −0.175, P = Japan, 2011 tion or more, of people engaging in prima- 0.031) and for males (β = −0.228, P= ry industry, and unemployment rate, annual 0.005), but not for females marriage rate, annual mean temperature, and annual postal savings per person Shiotsuki et al. 153 samples Annual total sunshine, annual mean 3.8 Lithium is inversely associated with 2016/Japan temperature, annual total rainfall, annual (SD ± 5.3; male but not female suicide after total snowfall 0.1–43) adjustment of meteorological factors Knudsen et al, 275 municipalities in No 0.6–30.7 Increasing suicide rate by increasing 2017 Denmark lithium levels Oliveira et al, 54 Portuguese Population density, income per capita, 10.88 No association between lithium in 2019 municipalities unemployment rates and proportion of (SD 27.18) public drinking water and suicide 2011–2016 Roman Catholics rates was found Palmer et al, Five drinking water The poverty had a parallel trend with 0.4–32.9 Significantly inverse associations for 2019 samples from each of suicide rate the overall SMRs and for males, but 15 Alabama counties not for females Giotakos et al. [22] reported lower suicide rates in prefectures with high levels of lithium in drinking water. Analyses were conducted with respect to lithium levels in 34 prefectures of Greece from both rural and urban areas. The results indicated that there was a tendency for lower suicide rates in the prefectures with high levels of lithium in drinking water. Confirmation of the association between levels of lithium in drinking water and suicide rate comes from a study published by Blüml et al. [27], which analysed 226 counties in Texas during 1999–2007. This study modeled the response of the county-level rate of suicide using both a linear and Poison rate regression adjusted for county-based population density, lithium levels, age, sex, race/ethnicity, median income per household, poverty and unemployment rates. Results showed that lithium levels were significantly associated with suicide rates. A recent study in Italy by Pompili et al. [98] found that lithium levels in tap water and local suicide rates were not significantly inversely related. Based on lithium levels and analysed suicide data at the community level, and including the following covariates: totally mountainous areas, highly urbanized, and geographic location, were reported that suicide rates were not statistically significantly correlated with lithium levels. The research group with Kapusta and colleagues [26] published a second study, Helbich et al. [25], on the same data set as that described previously, using a more refined statistical model based on the analysis of geospatial epidemiological data and lithium prescription. Previous studies have assumed that the lithium in drinking water originated from natural sources alone. They have not considered whether lithium prescribed to patients may have accumulated via waste water in groundwater aquifers after urinary excretion and interplay with natural lithium [23]. The results showed that lower lithium concentrations in the ground and drinking water might be responsible for higher suicides rates even after adjustment of lithium prescriptions. The most recently published study on the issue, Shiotsuki et al. [30], evaluated the association between lithium levels in the public water supply and suicide rates after adjustment of meteorological factors. The results suggest that trace lithium is inversely associated with male but not female suicide after adjustment of meteorological factors. Gender differences have been observed. Lithium decreases an impulsive aggressive behavior, which decreased [99]. It should be noted that findings of recent systematic review and meta- analysis [100] were obtained mostly from ecological studies which are not designed to make a causal relationship and decide on the individual level risk [101]. The only individual-level cohort study in this analysis was carried out by Knudsen et al. [97] that showed suicide insignificantly increases with

25 increasing exposure to lithium through drinking water in Denmark. There- fore, we need randomized, placebo-controlled trials for determining the definite effect of lithium intake through drinking water on suicide incidence, although these studies may be the subject of ethical and clinical challenges. Moreover, the cost-benefit analyses should be conducted to ensure that drinking water enrichment with lithium is both cost-effective and safe for vulnerable people such as children, pregnant women, and patients with special medical conditions. Due to the different patterns of suicide attempts and suicide mortality among men and women, evaluating the relationship between drinking water lithium and suicide attempts, as well as other violent actions and impulsive behaviors, may be fruitful. For example, in a systematic review and meta-analysis study Memon et al. [102] found that the protective (or inverse) association between lithium levels in drinking water and suicide mortality rates is likely to be stronger in populations with relatively higher suicide mortality rates.

1.3.2. Studies on potential anti-suicidal effects of lithium as a trace element in drinking water Another approach to determine lithium’s suicide protective effects is to investigate the ecological association between suicide rates and lithium in drinking water. Explication of the findings that even the very low levels of lithium provided in drinking water may reduce the risk of suicide is, at present, only speculative. It is possible, although unlikely, that the low levels of lithium such as those found in drinking water can bring about significant mood- stabilizing effects and reduce the risk of suicide in mood disorders not yet clinically defined by this mechanism [103]. In contrast, it could be speculated that even very low but long-lasting lithium exposure can enhance neurotrophic mechanisms, neuroprotective factors and/or neurogenesis, which may account eventually for a reduced risk of suicide [26, 104]. It has been estimated that although lithium probably prevents about 250 suicide deaths per year in Germany, only 0.06% of the German population were prescribed lithium [39]. Similarly, a UK study showed that at least 40% of patients with bipolar disorder who died by suicide were not prescribed lithium or a mood stabilizer [105]. Several researches have shown that low-dose lithium augments the efficacy of antidepressant medications [106]. For those who are suffering from mood disorders, including treatment-resistant depression, low-dose lithium supplementation may enhance the drug’s efficacy as well as reduce

26 the risk of side effects and drug toxicity associated with high-dose lithium treatment. Low dose lithium has also demonstrated an ability to prevent and delay relapse in people suffering from alcoholism as well as stabilizing and improving mood in drug users [18]. Lithium also exerts powerful antioxidant and anti-inflammatory effects, thus helping to counter neurodegenerative diseases involving inflammation and oxidative damage [107]. The result is a healthier brain and reduced risk of neurodegeneration and mood instability. While there is still a long way to go, and mental health still suffers more of a stigma than heart disease, for example, current research going on today, such as the mechanism by which lithium works, is an excellent step in the right direction [108]. It is an intriguing association, but not all articles are positive and the amount of exposure to lithium via drinking water is much, much lower than the exposure from lithium therapy. Estimates of the population attributable fraction suggested that 12% (95% CI 4%–20%) of suicide-related events could have been avoided if patients had taken lithium during the entire follow-up [109]. Overall, the findings summarized here, appear to provide strong and quite consistent support for the hypothesis that long term, but not acute, treatment with lithium may have a special role in reducing suicidal risks. It is important to acknowledge the significant role of mental disorders in suicide; underplaying it may result in missed opportunities for suicide prevention [110]. It seems we can step forward and conduct a series of clinical trials to test the hypothesis that long-term lithium intake through drinking water has a protective effect against suicide.

1.3.3. Summary of the literature review To summarize the literature review: in terms of the studies observed, evidence suggests that there is an inverse relationship between lithium levels in drinking water and suicide mortality, overall. Significance in most the outcome indicators were found in 14 of 42 studies, partial evidence was established in 2 of them (Pompili et al and Sugawara et al) [28, 98] and none in three piece of research (Knudsen et al, Oliveira et al, Kabacs et al) [93, 97, 111]. Unlike most international studies regarding natural lithium levels and suicide risk, no inverse relation was found in Portugal [111]. In summary, the suicide protective property of trace lithium was confirmed to have positive effect almost all studies. However, better understanding of how higher levels of lithium in drinking water can help in the prevention of suicide is required. Although lithium levels are extremely

27 low in drinking water, long-term exposure to lithium may be a factor which mitigates low absolute levels. Findings from a recent international multi-centre trial are compatible with the idea that long-term lithium treatment extends the survival of patients suffering from affective disorders to match the general population. Research from multiple countries, including the United States, has demonstrated a link between higher lithium levels in drinking water and lower rates of suicide in clinical and general populations [112]. It is still not clear what amount of lithium in drinking water can provide an independent protective effect for suicide. In particular, more research is needed to understand how lithium levels in the public drinking water supply correlate with blood lithium levels. Also, drinking water is not the only dietary source of lithium. According to the US Environment Protection Agency, some grains and vegetables are even richer in lithium than drinking water [92]. Overall, the findings summarized here, appear to provide strong and quite consistent support for the hypothesis that long term, but not acute treatment with lithium may have a special role in reducing suicidal risks. It is important to acknowledge the significant role of mental disorders in suicide; underplaying it may result in missed opportunities for suicide prevention [110]. Most evidence suggests that the negative association is present, but the heterogeneity of the studies and the lack of essential considerations in many of them make it hard to conclude such a connection is present. More needs to be done to shed light on an understanding of lithium and its effect in a low dose, as well.

28 2. MATERIAL AND METHODS

2.1. Study design

The research was designed as an ecological study using cross-sectional data gathered in municipalities (cities or district’s centres) of Lithuania. This study was performed at an aggregate level (no individual-level) to suggest an association between suicide mortality rate and lithium levels in drinking water. The study consists of two stages (Fig. 2.1.1).

Fig. 2.1.1. Study design overview *In study II, 1 drinking water sample was excluded from the statistical analyses because there exsisted very high lithium level (49.0 µg/L in wellfield of Šakiai district municipality). A single outlier was removed from the dataset (Annexe 3).

29 The unit of analysis was the municipality (district’s centres or cities). This ecological study was designed by aggregating suicide mortality rate (outcome) for each studied municipality for a five-year period and were analysed using linear regression with lithium levels in drinking water in the central wellfields of municipalities as exposure.

2.2. Methods

2.2.1. Study area: evaluation of lithium levels in drinking water For the analyses of lithium concentration in drinking water, we collected groundwater samples in the central wellfields of Lithuanian municipalities, which water supply systems serves the largest percentage of drinking water customers. Study I (Pilot study) Firstly, in the pilot study, we focused on the urban areas but not on the rural areas in order to decrease as much as possible the impact of the heterogeneity of social, economic and cultural background. In 2013 Lithuania had 2,971,905 inhabitants, 1,367,076 men and 1,604,828 women. There are 103 cities, 249 towns, and 21,000 villages in Lithuania [113] . The country’s 5 largest cities – Vilnius, Klaipėda, Kaunas, Šiauliai, Alytus (one exception was Panevėžys, because no response has been received from the department of wellfield) and 4 resort cities – Palanga, Neringa, Birštonas, Druskininkai had the population of 1,190,261 (41% of the total population of Lithuania, with a range from 2,525 in Birštonas to 526,356 in the capital city Vilnius [113]. This pilot study was conducted over a period of 3-month (November 2013–January 2014). Twenty-two samples (range from 1 to 5 per city) of drinking water from all (22) wellfields were taken in cities municipalities (5 samples – Vilnius, 4 samples – Kaunas, 3 samples – Klaipėda, 2 samples – Šiauliai, 2 samples – Alytus, 2 samples – Birštonas; 2 samples – Neringa, 1 sample – Druskininkai, and 1 sample – Palanga). The drinking water samples were analysed for lithium concentration by inductively coupled plasma mass spectrometry (ICP-MS) NexION 300D (Perkin Elmer, USA). The lithium detection limit, using seven replicated spiked with 1.0 µg/L of Multi-Element Calibration Standard 3 (Pure Plus PerkinElmer, USA) was 0.154 ± 0.03 µg/L and derived as described [114]. Study II The territory of Lithuania currently comprises 10 counties and 60 municipalities. The six largest municipalities by population (Vilnius, Kaunas,

30 Klaipėda, Šiauliai, Panevėžys and Alytus) are two different municipalities: municipality on the city and district. In the second study (Study II), over a period of 2-month (June 2017-July 2017), we collected 56 drinking water samples (1 sample per municipality, with one exception – 3 samples in Klaipėda municipality) from 56 central wellfields of the country’s municipalities, including cities municipalities (Druskininkai, Birštonas, Klaipėda, Neringa, Palanga, Kalvarija, Kazlų Rūda, Marijampolė, Rietavas, Visaginas, Elektrėnai) and districts municipalities (Alytus, Lazdijai, Varėna, Jonava, Kaišiadorys, Kaunas, Klaipėda, Kėdainiai, Prienai, Raseiniai, Klaipėda, Kretinga, Skuodas, Šilutė, Šakiai, Vilkaviškis, Biržai, Kupiškis, Panevėžys, Pasvalys, Rokiškis, Akmenė, Joniškis, Kelmė, Pakruojis, Radviliškis, Šiauliai, Jurbarkas, Šilalė, Tauragė, Mažeikiai, Plun- gė, Telšiai, Anykščiai, Ignalina, Molėtai, Utena, Zarasai, Šalčininkai, Širvin- tai, Švenčionys, Trakai, Ukmergė, Vilnius). In Klaipėda, city and district municipalities were taken 2 more drinking water samples (overall 3 samples in 3 wellfields) for assessment of different aquifers, which are exploited in drinking water supply systems. The mean value was calculated for analysis. The lithium concentration in these samples was determined by the ion chromatography DIONEX ICS-1000 (Thermo Scientific, USA) employing the standard LST EN ISO 14911. We cooperated with geologists and geoscientists from the Institute of Geology and Geography of Nature Research Centre in collecting drinking water samples and analysing the lithium concentration.

2.2.2. Dependent variable: suicide standardized mortality rate We obtained standardized mortality rate (SMR) for suicide (per 100,000 population) from the Health Information Centre of the Institute of Hygiene (Lithuania Database of Health Indicators) and used the average suicide SMR for 2009–2013 (for each of 9 municipalities) and 2012–2016 (for each of 53 municipalities). In accordance with International Statistical Classification of Diseases (ICD-10) the data included causes of death with codes X60-X84 [50, 115]. SMR were age standardized using the European Standard Population measure as defined by the World Health Organization [116].

2.2.3. Independent variables Municipality-based selected socio-demographic characteristics and morbidity indicators (incidence) that could be factors associated with suicide risk were examined. Municipal socio-demographic characteristics included unemployment rate (%), number of visits to psychiatrist per 100, divorce rate

31 per 1,000, women/men proportion (number of women per 1,000 men) and were obtained from the Department of Statistics, and from the Institute of Hygiene database, and were averaged across the investigated time period for each city or district’s municipality, respectively. In an addition to the municipal socio-demographic characteristics, morbidity indicators such as incidence (per 100,000) of mental and behavioral disorders (MBD) (ICD-10 codes F00-F99), incidence of diseases of the nervous systems (ICD-10 codes G00-99), number of suicide attempts (ICD-10 codes X60-84), and antidepressants usage rate across all age groups and gender within the five-year period (2012-2016) were included in the final analysis as an adjustment factors. Incidence of MBD included affective disorders (ICD-10 codes F30-39), schizophrenia (ICD-10 codes F20-29), MBD due to use of alcohol (ICD-10 codes F10). The analysis of antidepressant use was based on the Defined Daily Dose (DDD) that is most informative in general terms as respondent based methods create difficulty with estimating the level of use. The DDD based approach does not capture most aspects of clinical practice but provides an objective record of overall level of medicine use. Because wholesalers do not accumulate drug reserves, the DDD methodology provides the most adequate representation of drug use at the level of actual sales in the whole country. While the purchased drugs are not consumed entirely, there are no grounds to believe that failure of compliance at this level was different between demographically similar regions.

2.3. Statistical analysis

All statistical analyses were performed using software from the Statistical Package for Social Sciences 17.0 for Windows (SPSS, Inc., Chicago, Ill., USA). A value of P<0.05 was considered significant. A descriptive analysis was conducted to describe the profile of the municipalities. Tests for normality were conducted using a Kolmogorov- Smirnov test and a Pearson correlation coefficients or Spearman rank correlation coefficients were calculated between original and transformed variables of lithium and standardized suicide mortality rates, and among the municipalities’ routinely collected data. A curve estimation regression analysis was used in SPSS examining an exponential or quadratic relationship. Because of significant differences in population size across the municipalities, weighted least squares regression analysis, adjusted for the size of each population, was used to investigate the association between suicide SMRs (total, men, women) and lithium level in drinking water.

32 Study I (Pilot study) At first, we transformed a nonlinear model to a linear model, which can be analysed using Linear Regression procedure. When a nonlinear pattern was corrected by a log transformation, it was accompanied by a substantial increase in the value of R²: from 0.068 (P = 0.248) to 0.467 (P = 0.025). If the value of R² changed, then the relationship was not linear and the transformation was effective. The addition of the logarithmic term produced impressive results indicating that there is a strong relationship between suicide SMR and lithium levels, when lithium level is represented by a combination of the original variable and the log form of the original variable (R2 = 0.774, P = 0.005). Multivariate regression models incorporated the number of women for 1,000 men as covariate. Study II The least squares linear regression modelling weighted for local population size and including nonlinear terms into linear regression were performed. The most common non-linear term is the quadratic term constructing a regression model with the predictor and predictor2 terms. The quadratic term (X2) was included in the linear regression model because of the inverted U-shape relation of lithium to suicide SMR. To avoid a multicollinearity problem with the original variable [117] and its quadratic term (X2), a predictor variable (lithium) was centered (subtracting the mean) to create the square term: 2 2 Xc = (Lithium – mean) and Xc = (Lithium – mean) . The significance of quadratic terms could signal that the relationship is non-linear. The sign merely represents the type of non-linearity. A positive quadratic term could suggest that the relation is exponential. A negative relation suggests that for high values the relation becomes negative. The curve does not need to contain both sides of the U. A quadratic (squared) term turns a linear regression model into a curve. But because it is X that is squared, not the Beta coefficient, it still qualifies as a linear model (“linear regression” means linear in the Beta coefficients). The model with the quadratic term was proved to be significantly better. The formula for calculating the term is at x = –b/2a; following from y = –ax2+bx+c. When using the mean centered quadratic terms, the mean value should be added back to calculate the threshold turn value on the non-centered term x = –b/2a + mean (for purposes of interpretation when writing up results and findings).

33 We initiated a crude model analysis of the association of lithium levels in drinking water and suicide SMRs without any adjustment of the confounding factors. Please note the sign for x2 in each of the models. The sign is negative when the curve is concave (apex at the top, curve opens down). The associations of lithium levels in drinking water with suicide SMRs (total, men and women) were investigated adjusting for unemployment rate, visits to psychiatrists, divorce rate, and women/men proportion. Adjusted models include covariates with P<0.1 in a univariate analysis, apart from proportion of women per 1,000 men, which was correlated to population size. Analysis according to low and high lithium exposure groups Common statistical techniques multiple linear regression and ANCOVA were used and R-squared was used as the effect size. The mean lithium levels in drinking water were categorized into quartiles: Q1<3.5 μg/L, Q2 = 3.51–7.0 μg/L, Q3 = 7.1–20.0 μg/L, and Q4>20 to 39 μg/L. The municipalities were assigned to study groups on the basis of a median split. The median lithium concentration was calculated (7.0 μg/L) and municipalities with lithium concentration in drinking water greater than the median (N = 26) were assigned to the high lithium exposure group, the remaining municipalities (N = 27) were assigned to the low lithium exposure group. All analyses were repeated separately in these groups. We estimated the minimum sample sizes required for multiple linear regressions and ANCOVA by using Power and Sample Size software based on the pre-specified values of alpha, power and effect size (R2). A larger sample size is required for a smaller value of R-squared and when there are many explanatory (or independent) variables are to be tested in the regression model [118]. Sample size for a value of R-squared which is more than 0.4 the minimum sample size based on number of tested variables with selected R2 (Alpha = 0.05 and Power = 0.8) required will usually be less than 26 for a maximum of 5 independent variables [118]. Crude and multiple linear regression analysis were performed using the suicide SMRs (total, men, women) as the dependent variable. Multiple regression analyses were performed using forward or backward procedures, which first treated lithium levels as an independent variable, and then included municipalities’ characteristics as the independent variables. Because a bivariate correlation analysis of the independent variables showed potential for multicollinearity, Variance Inflation Factors (VIFs) were investigated. If all VIF scores were below the critical value of 5, as suggested by Fox (2002), multicollinearity was rejected [119]. The F-test with P<0.05 indicates that the final stepwise or backward regression model is statistically significant.

34 As the number of confounders was large, some may feature multicollinearity. Stepwise multiple regression analysis was used to build a regression model with minimum collinearity between the variables. Since the impact of individual confounders was of secondary interest, principal component analysis (PCA) was employed to obtain a reduced set of principal components (PC) that were uncorrelated and depicted the overall risk factor variability. Factor scores were generated using the Regression Method. The scatter-plots for visualization of relationships between variables were used. Multiple linear regressions were repeated with the reduced set of new variables (principal components) as the independent variables.

35 3. RESULTS

3.1. Lithium levels in drinking water, suicide SMRs, and socio-demographic characteristics in the municipalities

Study I (Pilot study) In total, 22 drinking water samples from the wellfields of 9 municipalities were analysed for lithium levels. Table 3.1.1 lists the municipalities’ socio- demographics characteristics, suicide SMR, and lithium level. Overall, for 5-year period (2009–2013), the average of suicide SMR in nine municipalities was 27 (range 16–50) per 100,000 of the population, 51 (range 29–93) per 100,000 of the population for men, and 7 (range 0–13) per 100,000 of the population for women. The average was 2.4 samples per municipality, with a range from 1 to 5 samples. The mean lithium level, as evidenced by raw values for 22 samples, was 10.9 (SD 9.1) μg/L ranged from 0.48 to 35.53 µg/L while median level was 3.6 µg/L.

Table 3.1.1. Suicide SMRs, lithium levels, and socio-demographic indicators in the municipalities (N = 9) Municipalities Population Women/ Suicide SMRb Lithium, a c in 2013 men Total Women Men µg/L Neringa 2,719 1,013 50.36 13.3 93.6 1.24 Klaipėda 158,541 1,205 19.33 5.7 36.3 13.1 Palanga 15,732 1,257 19.05 4.9 37.3 21.79 Šiauliai 106,470 1,304 26.88 9.2 50.1 28.68 Kaunas 306,888 1,269 21.03 9.3 36.1 11.63 Druskininkai 14,128 1,267 29.09 7.6 55.6 3.89 Birštonas 2,525 1,243 29.56 0 64.9 5.05 Vilnius 526,356 1,232 15.97 5.9 29.2 5.87 Alytus 57,281 1,177 29.7 7.2 56.8 6.94 a number of women for 1,000 men; b SMR, standardized mortality rate per 100,000 (European standard population); c average of lithium level.

There were municipal differences in suicide SMR of approximately 3.2 times for men (from 29.2 to 93.6 per 100,000) and approximately 13 times for women (from 0 to 13.3 per 100,000).

36 Study II The lithium levels in drinking water across 53 municipalities are shown in Fig. 3.1.2. The mean of lithium concentration across municipalities was 11.5 (SD 9.9) µg/L ranging from 1.0 to 39.0 µg/L, median – 7.0 (IQR 3.5– 20) µg/L.

Fig. 3.1.2. Lithium levels in drinking water samples collected from central wellfields of municipalities Six regions with different lithium levels in drinking water were distin- guished (Fig. 3.1.3). The Eastern Lithuania, Žemaitija and Šilutė had minimal values. While the regions of Central Lithuania, Northern Lithuania and Klaipėda had maximal values. These levels are obviously related to the aquifers of the corresponding lithological composition.

Fig. 3.1.3. Distribution of lithium levels in drinking water across Lithuania Descriptive statistics of suicide SMRs and municipal socio-demographic characteristics registered within the five-year period (2012–2016) are presented in Table 3.1.2.

Table 3.1.2. Suicide SMRs, lithium levels, and socio-demographic indicators in the municipalities (N = 53) Minimum Maximum Mean Std. Deviation Suicide SMR total 16.25 65.50 39.54 10.96 Suicide SMR men 31.89 137.09 79.05 22.17 Suicide SMR women 3.76 28.79 13.28 5.13 Unemployment rate, % 5.50 17.14 11.32 2.93 Divorce rate per 1,000 2.35 4.74 3.19 0.46 Visits to psychiatrist, n/100 7.27 55.14 27.46 8.17 Women/mena 972.40 1,275.40 1,135.87 47.85 SMR, standardized mortality rate per 100,000 (European standard population); a number of women for 1,000 men.

38 In addition, across municipalities, any registered incidence of MBD, affective disorders, schizoaffective disorders, diseases of nervous system, number of suicide attempts, antidepressants use, and MBD due to use of alcohol were included in the analysis (Table 3.1.3).

Table 3.1.3. Indicators of incidence, attempted suicide, and number of antidepressants use in the municipalities (N = 53) Incidences per 100,000 Minimum Maximum Mean Std. Deviation Affective disorder 149.62 991.47 474.43 226.68 Nervous system disease 1,019.11 8,762.92 4,424.28 1,554.20 Schizophrenia 14.77 110.91 49.65 23.403 MBD 1,457.27 4,194.89 2,893.79 655.72 Attempted suicide 13.30 108.90 52.21 24.36 Antidepressant use per 1,000a 1.00 35.8 19.97 8.30 MBD due to use of alcohol, total 86.23 528.66 286.41 98.35 MBD due to use of alcohol, women 27.74 215.01 116.01 45.31 MBD due to use of alcohol, men 160.56 901.37 477.35 173.89 MBD, mental and behavioral disorders; a number of the Defined Daily Dose/1,000 inhabitants/day.

3.2. Association of suicide SMRs with lithium levels in drinking water: Study I (Pilot study)

The pilot study analyses showed a nonlinear relationship between suicide SMRs and lithium levels in drinking water. We used the Curve Estimation procedure to identify functional relations between lithium levels and suicide SMRs. Firstly, we examined scatterplots that showed the pattern of both variables. The nonlinearity was assessed by including the fit line on the scatterplot. The logarithmic transformation was effective in reducing nonlinearity, both in terms of the increase in R² and the redistribution of points along both sides of the fit line (Fig. 3.2.1).

Fig. 3.2.1. Relationship between suicide SMRs and lithium levels Dotted line refers to a linear-linear regression and solid line to a linear-log regression model.

Pearson’s correlation analysis showed an inverse correlation between lithium level (natural log transformed), proportion of women/men, and suicide SMR in the total population and in men (Table 3.2.1). These variables did not significantly correlate with the suicide SMR in women.

Table 3.2.1. Associations of suicide SMRs with lithium levels, and proportion of women/men Pearson correlation coefficient (r) Variables Suicide SMR Total Men Women Lithium, µg/L –0.430 –0.016 –0.448 Log lithiuma –0.731* –0.728* –0.276 Women/menb –0.770* –0.726* -0.488 SMR, standardized mortality rate per 100,000 (European standard population); * p<0.05; a logged lithium level in drinking water; b number of women for 1,000 men.

40 In crude model, total suicide SMR (log transformed) were negatively associated with lithium levels (β = –0.91, P = 0.001). Regression models were weighted for local population size. After adjustment for proportion of women/1,000 men, the association with lithium level remained significant (β = –0.28, P = 0.034) in multiple regression analysis (Table 3.2.2).

Table 3.2.2. Association of suicide SMR with lithium levels Suicide SMR β t value P Model 1 Lithium, µg/L 0.560 1.789 0.117 Model 2 Log lithiuma –0.911 –5.841 0.001 Model 3 Log lithiuma –0.283 –2.886 0.034 Women/menb –0.713 –7.319 0.001 SMR, standardized mortality rate per 100,000 (European standard population); a logged lithium level; b number of women for 1,000 men.

Multiple regression analyses revealed that men suicide SMR were negatively associated with of log lithium (β = –0.96, P<0.001), but not for women SMR (β = 0.15, P = 0.70). Regression models were weighted for local population size. After adjustment for proportion of women/1,000 men, the association of suicide SMR with log lithium remain significant only in men (β = –0.70, P = 0.013) (Table 3.2.3).

Table 3.2.3. Association of suicide SMRs with lithium levels according to gender Suicide SMR Men Women β t value P β t value P Model 2 Log lithiuma –0.965 -9.684 <0.001 0.150 0.402 0.700 Model 3 Log lithiuma –0.702 –3.515 0.013 0.253 0.677 0.523 Women/menb –0.296 –1.484 0.188 0.490 1.311 0.238 SMR, standardized mortality rate per 100,000 (European standard population); a logged lithium level; b number of women for 1,000 men.

In summary, the pilot study demonstrated an inverse association between lithium level in drinking water (log natural transformed) and suicide SMR across 9 municipalities of Lithuania. Lithium levels in the drinking water were variable and ranged from 0.48 to 35.53 μg/L. Suicide SMR were a significantly inversely associated with lithium levels (log transformed) for total SMR (β = −0.28, P = 0.034) and for men (β = −0.70, P = 0.013), but not

41 for women (β = 0.15, P = 0.70) using weighted least squares regression analysis adjusted for the size of population.

3.3. Association of suicide SMR with lithium levels in drinking water: Study II

3.3.1. The correlates of suicide SMRs The distribution of lithium levels was considerably skewed (skewness = 0.948; kurtosis = –0.155). Thus, to use parametric statistical procedures, several transformations like logarithmic, square root and power (squared) were employed (Table 3.3.1.1).

Table 3.3.1.1. Correlation between suicide SMRs and lithium levels Pearson correlation coefficients (r) Original or transformed lithium level Suicide SMR Total Men Women Lithium, µg/L –0.150 –0.126 –0.153 Ln lithium –0.034 –0.210 –0.084 Quadratic term of lithium –0.253 –0.195 –0.229 Square root (Lithium, µg/L) –0.080 –0.076 –0.115 Centered: (Lithium – mean), µg/La –0.150 –0.126 –0.153 (Lithium – mean)2 –0.397* –0.286* –0.326* SMR, standardized mortality rate per 100,000 (European standard population); a Mean centering a variable means subtracting its (arithmetic) mean from all its values; *significant at P<0.05.

The total, men and women suicide SMRs were significantly and inversely associated only with quadratic term of centered lithium levels (Lithium level – mean)2. Non-parametric Spearman correlations (Table 3.3.1.2) show that suicide SMR is significantly correlated with the unemployment rate (r = 0.35; P = 0.010), population size (r = –0.32; P = 0.019) and the proportion of women/men (r = –0.38; P = 0.004).

42 Table 3.3.1.2. Correlation between suicide SMRs and socio-demographics indicators across municipalities by gender

Spearman correlation coefficient (rs) Suicide SMR Variables Total Men Women

rs P rs P rs P Unemployment rate, % 0.350 0.010 0.288 0.037 0.173 0.215 Visits to psychiatrists per 100 0.215 0.122 0.209 0.132 0.311 0.024 Divorces rate per 1,000 –0.169 0.240 –0.053 0.704 –0.001 0.992 Women/mena –0.384 0.004 –0.368 0.007 –0.217 0.119 Population size –0.322 0.019 –0.308 0.025 –0.162 0.245 SMR, standardized mortality rate per 100,000 (European standard population); a number of women for 1,000 men.

These three variables are thus potential confounders for influencing suicide mortality in a multivariate regression model. In contrast, the visits to psychiatrists correlated only with women suicide SMR (r = 0.31; P = 0.024) and the divorce rate did not significantly correlate with the suicide SMR.

3.3.2. Multiple regression analysis of suicide SMRs In the analysis to follow (Table 3.3.2.1–3.3.2.3), the suicide SMRs are the dependent variables. The data seem to show a combination of a positive linear and an inverted U-shaped quadratic trend. The linear component of how SMR changes as lithium level changes – is non-significant for all models. Divorces rate was not correlated with suicide SMR however it correlated with unemployment rate (r = 0.299, P = 0.012) and not included in Models 2. Proportion of women/men correlated with population size (r = 0.283, P = 0.040) and also not included in Models 2. Multiple regression analysis of total suicide SMRs Table 3.3.2.1 demonstrates multiple regression analysis of total suicide SMRs. In the quadratic equation of the crude model 1 the linear term is positive (β = 0.355, P = 0.070, non-significant coefficient) and the quadratic term is negative (β = –0.049, P = 0.007). In Model 2, the number of visits to psychiatrists and the unemployment rate were added as covariates. A negative quadratic term suggested that the expected direction and change of the suicide SMR – lithium slope changes as the value of X changes. Based on weighted least-squares regression modelling, lithium levels were significantly negatively associated with total suicide SMRs, only after it reached the level of 14.5 µg/L in multiple linear regression analysis (Table 3.3.2.1, Fig. 3.3.2.1).

43 Table 3.3.2.1. Association of total suicide SMRs with lithium levels Coefficientsa

Unstandardized Standardized P

B SE β Model 1 (Constant) 39.48 2.33

Xc [Lithium – mean] 0.355 0.192 0.320 0.070 2 Xc [Lithium – mean] –0.049 0.017 –0.482 0.007 Model: R2 = 0.175, F = 3.9, P = 0.027 Model 2 (Constant) 59.98 18.43

Xc [Lithium – mean] 0.280 0.173 0.252 0.112 2 Xc [Lithium – mean] –0.044 0.018 –0.436 0.018 Unemployment rate, % 0.876 0.353 1.220 0.017 Visits to psychiatrists per 100 0.482 0.173 0.338 0.008 Model: R2 = 0.338, F = 6.3, P<0.001 SMR, standardized mortality rate per 100,000 (European standard population); SE, Standard error; a Weighted Least Squares Regression – Weighted for population size; Model 2: Adjusted for unemployment rate and number of visits to psychiatrist.

The graphical evidence hints at the curvilinear relationship between lithium levels in drinking water and total suicide SMR (Fig. 3.3.2.1). Notably, the regression findings presented in Table 3.3.2.1 largely confirm such a relationship. This table contains the estimation results of the regression equation (y = –ax2 + bx + c), where ‘a’ and ‘b’ capture the first-order and the second-order effect of lithium on suicide SMR.

Fig. 3.3.2.1. A scatter-gram of the lithium levels in drinking water versus total suicide SMR Multiple regression analysis of men suicide SMR Again, linear and quadratic regression models were fitted to identify areas that might show linear or inverted U-shaped relationships between lithium levels and men suicide SMR (Fig. 3.3.2.2).

Fig. 3.3.2.2. A scatter-gram of the lithium levels in drinking water versus men suicide SMR

45 The inverse association of men suicide SMR with lithium concentration in drinking water was on the margin of significance (P = 0.055) only after it reached the level of 15.3 µg/L in multiple linear regression analysis (Table 3.3.2.2, Fig. 3.3.2.2).

Table 3.3.2.2. Association of men suicide SMRs and lithium levels Coefficientsa Unstandardized Standardized B SE β P Model 1 (Constant) 76.6 4.8

Xc [Lithium – mean] 0.702 0.173 0.252 0.086 2 Xc [Lithium – mean] –0.075 0.036 –0.365 0.046 Model: R2 = 0.083, F = 2.2, P = 0.115 Model 2

Xc [Lithium – mean] 0.554 0.351 0.250 0.118 2 Xc [Lithium – mean] –0.073 0.037 –0.355 0.055 Unemployment rate, % 2.009 0.746 1.379 0.010 Visits to psychiatrists per 100 0.942 0.366 0.325 0.013 Model: R2 = 0.281, F = 5.07, P = 0.001 SMR, standardized mortality rate per 100,000 (European standard population); SE, Standard error; a Weighted Least Squares Regression – Weighted for populations size; Model 2: adjusted for unemployment rate and number of visits to a psychiatrist.

Multiple regression analysis of women suicide SMRs Based on weighted least-squares regression modelling, lithium concentration was significantly negatively associated (P = 0.040) with women suicide SMR, only after it reached the level of 13.2 µg/L in multiple linear regression analysis (Fig. 3.3.2.3 and Table 3.3.2.3). Therefore, lithium in drinking water adjusted for unemployment rate and number of visits to a psychiatrist may reduce suicide SMR.

Fig. 3.3.2.3. A scatter-gram of the lithium levels in drinking water versus women suicide SMR

Table 3.3.2.3. Association of women suicide SMRs with lithium levels Coefficientsa Unstandardized Standardized B SE β P Model 1 (Constant) 13.5 0.95 <0.001 Xc [Lithium – mean] 0.093 0.079 0.206 0.244 Xc2 [Lithium – mean] –0.017 0.007 –0.424 0.019 Model: R2 = 0.109, F = 3.1, P = 0.056 Model 2 (Constant) 18.5 8.1 0.025 Xc [Lithium – mean] 0.058 0.075 0.128 0.449 Xc2 [Lithium – mean] –0.017 0.008 –0.407 0.040 Unemployment rate, % 0.256 0.154 0.882 0.103 Visits to psychiatrists per 100 0.207 0.076 0.359 0.009 Model: R2 = 0.227, F = 4.1, P = 0.004 SMR, standardized mortality rate per 100,000 (European standard population); SE, Standard error; a Weighted Least Squares Regression – Weighted for population size; Model 2: adjusted for unemployment rate and number of visits to psychiatrist.

47 In summary, when considering the effect of lithium intake with drinking water associated with a lower suicide SMR in a nonlinear way; an anti- suicidal effect of lithium in drinking water is not present if lithium level is below a certain level. That is, a function of lithium level has both increasing and decreasing part. Lithium levels in drinking water were significantly negatively associated with the total suicide SMR only after it reached the certain level.

3.4. Relationships between suicide SMRs, incidence of affective disorders, and lithium levels

3.4.1. Analysis of suicide SMRs and other indicators of municipalities according to quartiles of lithium levels In the study above we found that lithium in drinking water, adjusted for unemployment rate and number of visits to a psychiatrist, may reduce suicide SMR where the lithium level in drinking water was above a certain level. In current analysis, the mean lithium levels were categorized into quartiles (<3.5, 3.51–7.0, 7.1–20.0, and >20 to 39 μg/L) and into two groups according to a median of 7.0 μg/L, i.e. according to the quartiles Q1–Q2 (the low lithium level exposure group) and Q3–Q4 (the high lithium level exposure group). Examined variables including suicide SMR and incidence of MBDs, schizophrenia, and nervous system disease were then calculated according to the quartiles of lithium level. As shown in Table 3.4.1.1, we found no clear differences in suicide SMR (total, men and women) in relation to the quartiles of lithium level. ANOVA testing revealed significant differences in incidence of MBD (F3,49 = 6.79, P = 0.001) and MBD due to use of alcohol for women (F3,49 = 6.23, P = 0.001) between the quartiles of lithium level. Increased incidence of MBD due to use of alcohol for women was observed for the three upper quartiles of lithium level compared with the lowest quartile. Incidence of affective disorders in the highest quartile was higher than in the lowest quartile of lithium levels in drinking water with a clear tendency to significance (F3,49 = 2.74, P = 0.053).

48 Table 3.4.1.1. Differences of suicide SMRs and incidence indicators according to quartiles of lithium levels Lithium quartiles, µg/L Variables, F 3,49; P per 100,000 Q1 Q2 Q3 Q4 <3.5 3.5–7.0 7.1–20 >20 Suicide SMR total 41.76± 43.04± 45.53± 39.3± F = 0.65 13.64 9.03 10.29 13.03 P = 0.587 Suicide SMR men 78.38± 78.37± 84.63± 74.07± F = 0.49 29.66 20.42 17.16 21.31 P = 0.689 Suicide SMR women 13.88± 12.47± 13.65± 13.28± F = 0.19 4.16 4.76 4.62 7.23 P = 0.901 Affective disorder 395.40± 391.32± 531.82± 590.07± F = 2.74 156.504 221.67 229.96 246.301 P = 0.053 MBD 2,470.62± 2,631.24± 3,281.26± 3,206.49± F = 6.79 669.813,4 604.813,4 518.411,2 655.721,2 P = 0.001 MBD due to use 223.52± 307.61± 303.67± 309.65± F = 2.56 of alcohol, total 81.902,3,4 118.601 71.941 97.261 P = 0.065 MBD due to use 76.19± 119.20± 136.31± 131.69± F = 6.23 of alcohol, women 28.48 42.99 38.86 46.611,2,3 P = 0.001 MBD due to use 389.93± 515.39± 490.84± 511.93± F = 1.55 of alcohol, men 158.43 209.07 138.92 169.37 P = 0.213 Schizophrenia 47.2± 42.53± 60.08± 47.97± F = 1.44 22.95 21.44 26.62 20.54 P = 0.240 Nervous system 4,195.45± 4,521.45± 5,013.44± 3,871.45± F = 1.31 disease 1,969.94 1,676.53 1,326.52 966.96 P = 0.280 Data are presented as mean ± SD; SMR, standardized mortality rate per 100,000 (European standard population); MBD, mental and behavioral disorders; 1,2,3,4 the number in superscript indicates the quartiles of the lithium level in drinking water, with significant differences P<0.05 (post hoc with Bonferroni correction).

However, there were a series of factors that could make significant contributions to risk of suicidal behavior such as incidence of diseases of nervous system and schizophrenia. The distribution of means of these disorders did not differ significantly across quartiles of lithium.

3.4.2. Analysis of suicide SMRs and other indicators of municipalities according to a median of lithium levels The median concentration for lithium in drinking water was 7.0 μg/L. All analyses were repeated separately for the two groups: low (27 municipalities) and high (26 municipalities) lithium exposure groups, categorized to median of lithium level. The descriptive characteristics of the groups are presented in Table 3.4.2.1.

49 There were no significant differences in socio-demographic characteristics of the municipalities between the two groups categorized to median of lithium level (Table 3.4.2.1). Table 3.4.2.1 indicates that distribution of the affective disorders (F1,51 = 20.26, P<0.0001) and the MBDs (F1,51 = 8.0, P = 0.007) were increased in the high lithium exposure group compared with the low lithium exposure group. In the total sample the antidepressants usage rate was significantly positive correlated with lithium levels (r = 0.290, P = 0.035) and tended to be positively associated with suicide SMR (r = 0.254, P = 0.067) (data not shown). The antidepressant usage rate correlated with incidence of affective disorders (r = 0.548, P<0.001) and incidence of mental and behavioral due to use of alcohol (r = 0.362, P = 0.008). Antidepressants usage tended to be higher in high lithium exposure group compared with the low lithium exposure group (22.14±7.42 and 17.89±8.61; F 1,51 = 3.64, P = 0.062) (Table 3.4.2.1). The relative difference of incidence of affective disorders was 26.9% and the relative difference of antidepressants use was 19.2% between the low and high lithium exposure groups. Attempted suicide number marginally correlated with total suicide SMR only in the high lithium exposure group (r = 0.395, P = 0.046). Distribution of suicide attempts was equal between lithium exposure groups (F1,51 = 0.006, P = 0.938). In the total sample number of attempted suicides tended to correlate with number of visits to psychiatrists (r = 0.248, P = 0.073) and was negatively associated with incidence of diseases of the nervous system (r = –0.287, P = 0.037) (data not shown). In addition, an increased incidence of MBD due to use of alcohol in women was found in the high lithium exposure group compared with the low lithium exposure group (F1,51 = 9.57, P = 0.003). The incidence of diseases of the nervous system and schizophrenia did not differ between the lithium exposure groups (Table 3.4.2.1).

50 Table 3.4.2.1. Differences of suicide SMRs and sociodemographic indicators according to the lithium exposure group Low lithium High lithium Variables, Overall, F exposure group exposure group P per 100,000 n = 53 (1,51) N = 27 N = 26 Suicide SMR 42.42±11.27 42.67±11.8 42.54±11.43 0.006 0.938 total Suicide SMR 78.37±24.79 79.75±19.54 79.05±22.17 0.05 0.824 men Suicide SMR 13.15±4.45 13.41±5.85 13.28±5.13 0.03 0.854 women Populations size 42176±64932 36418±21224 39351±48302 0.156b Median 26002.3 34205.8 28328.9 Women/mena 1137.07±49.52 1134.62±46.99 1135.87±47.85 0.34 0.854 Unemployment 11.18±3.29 11.46±2.56 11.32±2.93 0.13 0.724 rate, % Divorces rate per 3.12±0.52 3.26±0.37 3.19±0.46 1.28 0.262 1,000 Visits to psy- 26.86±8.49 28.07±7.94 27.46±8.17 0.29 0.594 chiatrists per 100 MBD 2553.90±629.84 3246.75±476.91 2893.79±655.72 20.26 <0.001 Affective 393.28±189.41 558.71±234.66 474.43±226.68 8.0 0.007 disorder total men 205.41±123.64 278.85±141.92 242.13±136.94 4.11 0.048 women 564.64±253.90 783.12±324.99 673.12±309.19 7.57 0.001 Nervous system 4364.49±1795.55 4486.37±1289.76 4424.28±1554.20 0.08 0.778 disease Schizophrenia 44.98±21.90 54.49±24.33 49.65±23.40 2.24 0.141 MBD due to use 267.12±109.37 306.43±82.84 286.41±98.35 2.16 0.148 of alcohol, total MBD due to use 454.99±193.70 500.57±150.90 477.35±173.89 0.91 0.345 of alcohol, men MBD due to use 98.49±42.16 134.18±41.79 116.00±45.31 9.57 0.003 of alcohol, women Attempted suicide 57.50±4.75 54.88±4.82 52.21±24.36 0.006 0.938 Antidepressant 17.89±8.61 22.14±7.42 19.97±8.30 3.64 0.062 use per 1,000* SMR, standardized mortality rate per 100,000 (European standard population); MBD, mental and behavioral disorders; SD, standard deviation; a number of women for 1,000 men; b Mann- Whitney test; *number of the Defined Daily Dose /1,000 inhabitants/day.

51 In Fig. 3.4.2.1 and Fig. 3.4.2.2 presented scatterplots of association between lithium and incidence of affective disorder and suicide SMR for both lithium exposure groups. According to Fig. 3.4.2.1 (at right), the fit line approaches a plateau (with a slight increase) in incidence of affective disorders in municipalities with lithium level higher than median (7.0 µg/L). The relative difference of incidence of affective disorders was 26.9% between the low and high lithium exposure groups.

Fig. 3.4.2.1. Association of incidence of affective disorders with lithium level, linear (left, P = 0.004) and cubic (right, P = 0.020) Dash line indicate median: horizontal, of the total incidence of affective disorders 419/100,000; vertical, of the lithium level in drinking water 7.0 µg/L.

Fig. 3.4.2.2. Association of suicide SMRs with lithium level, linear (left, P = 0.212) and cubic (right, P = 0.030) Vertical dash line indicates median of the Lithium level in drinking water 7.0 µg/L.

The results suggest a higher incidence of affective disorders in municipalities above median of lithium level in drinking water compared with other

52 municipalities with the same socio-demographic and psychiatric character- ristics. In these municipalities suicide SMR was inversely associated with lithium level in drinking water (Fig. 3.4.2.2, in right). Thus, the presence of affective disorders could be an important condition for the linked between lithium level and suicide mortality. The bivariate relationships between all variables were calculated by Spearman correlation coefficients and a correlation matrix for low and high lithium exposure groups was presented in Annex 1 and Annex 2.

3.4.3. Associations of suicide SMRs and mental and behavioral disorders with lithium levels in low lithium exposure group Table in Annex 1 demonstrates the correlation between variables for the low lithium exposure group (below the median of 7.0 µg/L). We evaluated how closely lithium, suicide SMR, socio-demographic variables and different incidence of psychiatric disorder were associated. A high correlation exists between the total suicide SMR and the suicide SMR for men with r = 0.92 (P<0.011). However, the total SMR is lower when correlated with women SMR (r = 0.48, P<0.05) and the correlation between men and women is even lower (r = 0.39, P<0.05). We found that the lithium level in drinking water was not associated with suicide SMR in the low lithium exposure group. However, a positive correlation between lithium level and incidence of MBD due to use of alcohol for total and for men and women there was found. The incidence of MBD due to use of alcohol (total, for men and women) had a positive correlation with incidence of MBDs (r = 0.49, P<0.01; r = 0.48, P<0.05; r = 0.48, P<0.05 respectively). The incidence of MBD due to use of alcohol for women had a strong association with incidence of MBD due to use of alcohol for men (r = 0.68, P<0.01). Incidence of MBDs were more strongly associated with incidence of affective disorders (r = 0.71, P<0.01) than with incidence diseases of the nervous system (r = 0.39, P<0.05). Multiple regression analysis for the suicide SMR Total suicide SMR were not associated with lithium levels in the crude model (β = 0.141, P = 0.713). In the adjusted model 2 (Table 3.4.3.1), total suicide SMR was also not associated with lithium levels in drinking water. However, total suicide SMR was associated with unemployment rate (β = 0.494, P = 0.003) and incidence of affective disorder (β = 0.372, P = 0.022). These variables explained over half (52.8%, P<0.001) of variation in total suicide SMR.

53 Table 3.4.3.1. Association of suicide SMRs with lithium levels in the low lithium exposure group Coefficients

Unstandardized Standardized t P

B SE β Model 1 Suicide SMR total (Constant) 39.37 4.81 8.18 <0.001 Lithium, µg/L 0.189 1.08 0.141 0.713 0.482 Model: R2 = 0.001, F = 0.029, P = 0.867 Model 2 Suicide SMR total (Constant) 6.112 5.981 1.022 0.317 Unemployment rate, % 1.969 0.608 0.494 3.241 0.003 Affective disorders 0.029 0.012 0.372 2.442 0.022 Model: R2 = 0.528, F = 13.42, P<0.001 Model 3 Suicide SMR women (Constant) 0.925 2.328 0.397 0.695 Affective disorders 0.014 0.004 0.503 3.443 0.002 Visits to psychiatrist per 100 0.246 0.086 0.416 2.846 0.009 Model: R2 = 0.509, F = 12.43, P<0.001 Model 4 Suicide SMR men (Constant) -14.434 16.854 -0.856 0.400 Unemployment rate, % 3.512 1.278 0.439 2.748 0.011 MBD 0.018 0.007 0.397 2.489 0.020 Model: R2 = 0.498, F = 11.90, P<0.001 SMR, standardized mortality rate per 100,000 (European standard population); MBD, mental and behavioral disorders; SE, standard error; Model 1, crude model; Models 2, 3, 4, adjusted for baseline characteristics (local population size, proportion of women/men, unemployment rate, divorce rate, visits to psychiatrists), and following adjustment for the incidence of the MBD, affective disorders, MBD due to use of alcohol, schizophrenia, and diseases of nervous system.

54 There were significant and positive associations between unemployment rate (β = 0.439, P = 0.011) and incidence of MBD (β = 0.397, P = 0.020) and suicide SMRs among men (Model 4; R2 = 0.498, P<0.001), as well as number of visits to a psychiatrist (β = 0.416, P = 0.009) and incidence of affective disorder (β = 0.503, P = 0.002) and suicide SMRs among women (Model 3; R2 = 0.509, P<0.001) (Table 3.4.3.1). The curves for visualization of the association of incidence of affective disorders and total suicide SMR with lithium level below the median <7.0 µg/L) displayed in Fig. 3.4.3.1.

Fig. 3.4.3.1. Associations of incidence of affective disorders (left) and total suicide SMRs (right) with lithium level in the low lithium exposure group (below the median of 7.0 µg/L)

3.4.4. Relationships between suicide SMRs, mental and behavioral disorders and lithium levels in high lithium exposure group Spearman correlations were calculated between the total SMR, SMRs by gender and selected socio-demographic variables, the incidence of MBD disorders and diseases of the nervous system associated with high suicide SMRs in municipalities with high lithium exposure (above the median of 7.0 µg/L) (Table 3.4.4.1 and Table 3.4.4.2: full correlation matrix table presented in Annex 2). A high correlation existed between the total suicide SMR and the suicide SMR for men with r = 0.93 (P<0.001) (Table 3.4.4.1). However, the total suicide SMR was lower when correlated with women SMR (r = 0.67, P<0.01) and the correlation between men and women was even lower (r = 0.52,

55 P<0.01). Unemployment rate showed numerous significant positive correlations, with total suicide SMR while population size was negatively associated with suicide SMR in total (rs = –0.47; P<0.01) and in men (rs = –0.39; P<0.01).

Table 3.4.4.1. Correlation between suicide SMRs and socio-demographic indicators in the high lithium exposure group

Spearman correlation (rs) Variablesa 1 2 3 4 5 7 1 Lithium, µg/L 1 2 Suicide SMR total –0.46* 1 3 Suicide SMR men –0.45* 0.93** 1 4 Suicide SMR women –0.34 0.67** 0.52** 1 5 Populations size 0.21 –0.47** –0.39* –0.36 1 6 Women / menb 0.42* –0.32 –0.23 –0.30 0.06 7 Unemployment rate, % –0.24 0.50** 0.41* 0.47* –0.43* 1 17 Antidepressant usec 0.29* 0.25 –0.02 0.07 –0.23 0.228 SMR, standardized mortality rate per 100,000 (European standard population); a the numbering of the variables is conditional; b number of women for 1,000 men; c number of the Defined Daily Dose /1,000 inhabitants/day; *P<0.05; **P<0.01.

Table 3.4.4.2. Correlation between suicide SMRs, socio-demographic and incidence indicators in the high lithium exposure group

Spearman correlation (rs) Variablesa 3 8 9 10 11 13 14 15 3 Suicide SMR 1 men 8 Divorces rate 0.09 1 per 1,000 9 Visits to psy- 0.14 0.16 1 chiatrists per 100 10 Affective –0.19 –0.02 0.09 1 disorder 11 Nervous system 0.07 0.41* –0.19 0.43* 1 disease 12 Schizophrenia 0.19 0.22 0.43* 0.38 0.44* 13 MBD –0.21 0.06 –0.06 0.63** 0.40* 1 14 MBD due to 0.26 0.26 0.29 –0.29 –0.18 0.21 1 use of alcohol, total

56 Table 3.4.4.2. Continued

Spearman correlation (rs) Variablesa 3 8 9 10 11 13 14 15 15 MBD due to 0.20 0.28 0.23 –0.24 –0.13 0.28 0.96** 1 use of alcohol, men 16 MBD due to 0.42* 0.31 0.41* –0.17 0.07 0.22 0.80** 0.71** use of alcohol, women 17 Antidepressant –0.02 0.05 0.46* 0.59*** 0.07 0.53*** 0.32* 0.35* useb SMR, standardized mortality rate per 100,000 (European standard population); MBD, mental and behavioral disorders; a the numbering of the variables is conditional; b number of the Defined Daily Dose /1,000 inhabitants/day; *P<0.05;**P<0.01.

The curves for visualization of the associations of incidence of affective disorders and suicide SMRs with lithium level above the median (≥7.0 µg/L) displayed significant suicide curve decrements and invers direction in affective disorders curve (Fig. 3.4.4.1).

Fig. 3.4.4.1. Associations of incidence of affective disorders (left) and suicide SMRs (right) with lithium levels in the high lithium exposure group (above the median of 7.0 µg/L)

57 Tables 3.4.4.3–3.4.4.5 demonstrate the results of the multiple linear regression analysis of the suicide SMR.

Table 3.4.4.3. Association of total suicide SMR with lithium level in the high lithium exposure group Coefficients Variables Unstandardized Standardized t P B SE β Crude model (Constant) 55.181 5.325 10.363 <0.001 Lithium, µg/L –0.643 0.252 –0.462 –2.555 0.017 Model: R2 = 0.462; F = 6.529, P = 0.017 Adjusted modela (Constant) 30.679 11.178 2.745 0.012 Lithium, µg/L –0.504 0.236 –0.363 –2.135 0.044 Unemployment rate, % 1.901 0.781 0.413 2.433 0.023 Model: R2 = 0.375, F = 6.894, P = 0.005 SE, standard error; a adjusted for baseline characteristics (local population size, proportion of women/men, unemployment rate, divorce rate, visits to psychiatrists per 100), and following adjustment for the incidence of the MBD, affective disorders, MBD due to use of alcohol, MBD due to use of alcohol women, MBD due to use of alcohol men, diseases of nervous system, schizophrenia.

In unadjusted crude model (Table 3.4.4.3), the results reveal that lithium (β = –0.462, P = 0.017) was significant predictors of the suicide SMR (R2 = 0.462). In adjusted model 2, the results reveal that lithium (β=–0.363, P = 0.044) and unemployment rate (β = 0.413, P = 0.023) were all significant predictors of the suicide SMR (R2 = 0.375, P = 0.005). In this model, the maximum value of VIF was 1.252, which did not exceed 5, and the minimum value of tolerance was 0.942 indicating no multicollinearity. In unadjusted crude model (Table 3.4.4.4), the results revealed that lithium level (β = –0.343, P = 0.087) was not significant predictors of women suicide SMR. However, after adjustment the result revealed that lithium level (β = –0.455, P = 0.015), incidence of disease of the nervous system (β = –0.560, P = 0.007) and incidence of schizophrenia (β = –0.5, P = 0.011), were significant predictors of women suicide SMR (R2 = 0.423, P = 0.006). In this model, the maximum value of VIF was 1.322, which did not exceed 5, and the minimum value of tolerance was 0.751 indicating no multicollinearity.

58 Table 3.4.4.4. Association of women suicide SMR with lithium level in the high lithium exposure group Coefficients Variables Unstandardized Standardized t P B SE β Crude model (Constant) 18.008 2.795 6.444 <0.001 Lithium, µg/L –0.236 0.132 –0.343 –1.787 0.087 Model: R2 = 0.117, F = 3.194, P = 0.087 Adjusted modela (Constant) 24.215 4.744 5.104 <0.001 Lithium, µg/L –0.306 0.116 –0.455 –2.637 0.015 Nervous system disease –0.003 0.001 –0.560 –2.993 0.007 Schizophrenia 0.120 0.043 0.500 2.767 0.011 Model: R2 = 0.423, F = 5.369, P = 0.006 SE, standard error; a adjusted for baseline characteristics (local population size, proportion of women/men, unemployment rate, divorce rate, visits to psychiatrists), and following adjustment for the incidence of the MBD, affective disorders, MBD due to use of alcohol, MBD due to use of alcohol women, MBD due to use of alcohol men, schizophrenia, and diseases of nervous system.

There was a crude negative association (Table 3.4.4.5) between the men suicide SMR and lithium level (β = –0.452, t = –2.484, P = 0.020; R2 = 0.204). Adjusting for socio-demographic variables was no impacted considerably on the association, but adding incidence of the MBD reinforced the association. In full adjusted model for men, the results revealed that lithium (β = –0.465, P = 0.009) and MBD due to use of alcohol for women (β = 0.433, P = 0.014), were significant associated with men suicide SMR (R2 = 0.391, P = 0.003)

Table 3.4.4.5. Association of the men suicide SMR with lithium level in the high lithium exposure group Coefficients Variables Unstandardized Standardized t P B SE β Crude model (Constant) 99.988 8.863 11.281 <0.001 Lithium, µg/L, –1.040 0.419 –0.452 –2.484 0.020 Model: R2 = 0.204, F = 6.168, P = 0.020

59 Table 3.4.4.5. Continued Coefficients Variables Unstandardized Standardized t P B SE β Adjusted modela (Constant) 73.394 12.758 5.753 <0.001 Lithium, µg/L –1.068 0.374 –0.465 –2.855 0.009 MBD due to use of alcohol, 0.202 0.076 0.433 2.659 0.014 women Model: R2 = 0.391, F = 7.398, P = 0.003 SE, standard error; a adjusted for baseline characteristics (local population size, proportion of women/men, unemployment rate, divorce rate, visits to psychiatrists), and following adjustment for the incidence of the MBD, affective disorders, MBD due to use of alcohol, MBD due to use of alcohol women, MBD due to use of alcohol men, schizophrenia, and diseases of nervous system.

Findings were used to develop and interpret likely models of lithium in drinking water, unemployment, incidence of MBD and suicide SMR. Suicide SMR was associated with lithium in drinking water in the high lithium exposure group. Multiple regression analysis revealed that the total suicide SMR was negatively associated with lithium in drinking water and significant positive relationships were found between the proportions of unemployment. There are different factors associated with suicide depending on gender, which may also point to different psychological mechanisms in the background of suicide. The highest local R2 values were found in the suicide SMR for women explained approximately 42% of the variation in the suicide SMR. An incidence of MBD due to use of alcohol in women appears to cause increased risk of suicidal behavior in men. The incidence of MBD was strongly related to suicide SMR. MBD due to use of alcohol in women was the most important factor suicide SMR for men.

3.4.5. Exploratory factor analysis of the variables in high lithium exposure group: supplementary analysis In supplementary analysis we used factor analysis, for data reduction to reduce the number of multiple comparisons in regression analysis of suicide SMRs (as has been analyzed above in chapter 3.4.4). In present study we use suicide SMRs as dependent variable and both lithium level in drinking water

60 and unemployment rate as independent variables. Using a principle components analysis followed by a varimax rotation, we identified factor structures for 11 potentially confounding characteristics of municipalities. The theore- tical basis for factor analysis is that variables are correlated because they share one or more common components. Table 3.4.5.1 summarizes factor analyses of 11 variables in the high lithium exposure group. It could be seen from Table 3.4.5.1, that out of 11 municipalities variables four factors have been extracted and this four factors, together explain the total variance of this municipal variables to the extent of 76.46%. The incidence of affective or mental and behavioral disorders exists in combination with sociodemographic characteristics. Total variance explained in the 4-component PCA as 70.87%.

Table 3.4.5.1. The exploratory factor analysis of the variables in the high lithium exposure group Component (the loading)

PC1 PC2 PC3 PC4 MBD due to use of alcohol, total 0.976 MBD due to use of alcohol, men 0.921 MBD due to use of alcohol, women 0.831 Women/mena 0.534 MBD 0.850 Affective disorders 0.806 Population size –0.643 Disease of nervous system 0.819 Divorces rate per 1,000 0.731 Visits to psychiatrist per 100 0.762 Schizophrenia 0.532 Eigenvalue 3.21 2.51 1.46 1.23 % of variation explained 27.86 20.82 14.22 13.54 Cumulative % variation explained 27.86 48.69 62.91 76.46 MBD, mental and behavioral disorders; a number of women for 1,000 men; Loadings > 0.5 are included; eigenvalue >1 rule; Kaiser-Meyer-Olkin test – 0.576; Bartlett's Test – 145.01, df=55, P<0.001.

In multivariate regression analysis of suicide SMRs (total, men, and women) were included lithium and unemployment rate as independent variables following by four components PC1–PC4 as potentially confounding variables. The models created fitted the data well: for total SMR R2 = 0.375,

61 P = 0.005; for women R2 = 0.603, P<0.001; for men R2 = 0.204, P = 0.020 (Table 3.4.5.2).

Table 3.4.5.2. Associations between suicide SMRs and lithium level in the high lithium exposure group Coefficients Unstandardized Standardized t P B SE β Model 1 (backward) Suicide SMR total (Constant) 30.579 11.168 2.735 0.013 Lithium, µg/L –0.503 0.246 –0.363 –2.134 0.044 Unemployment rate, % 1.931 0.881 0.413 2.432 0.023 Model: R2 = 0.375, F = 6.894, P = 0.005 Model 2 (backward) Suicide SMR women (Constant) 21.141 2.051 10.319 <0.001 Lithium, µg/L –0.387 0.098 –0.576 –2.482 0.001 PC3 – Nervous system diseases –3.077 0.817 –0.537 –3.784 0.001 PC4 – Schizophrenia 2.906 0.821 0.541 3.739 0.012 Model: R2 = 0.603, F = 11.142, P<0.001 Model 3 (stepwise) Suicide SMR men (Constant) 99.988 8.763 11.281 <0.001 Lithium, µg/L –1.041 0.427 –0.452 –2.484 0.021 Model: R2 = 0.204, F = 6.168, P = 0.020 SMR, standardized mortality rate per 100,000 (European standard population); SE, standard error; All models (for total, women and men suicide SMR) were adjusted for unemployment rate controlling for principal components PC1–4.

In summary, the supplementary analyses showed the same results as the main multiple regression analysis (above in chapter 3.4.4). In present ecological study, we evaluated the associations between suicide SMRs and the lithium levels adjusted by unemployment rate and by confounders – principal components PC1–4 in which the incidence of affective or mental and behavioral disorders exists in combination with sociodemographic variables of municipalities. In the adjusted models, total, men and women suicide SMRs were also associated with lithium level in drinking water.

62 4. DISCUSSION

4.1. The lithium level in drinking water of the central wellfields in Lithuanian municipalities

Lithium level from central drinking water systems across Lithuania demonstrated a geographical distribution across the municipalities. Naturally occurring lithium is not systematically monitored in Lithuanian drinking water. According to our study results, the mean lithium concentration of fifty- three district municipalities in the public drinking water samples was 11.5 (SD 9.9) µg/L. The geographic variation of lithium levels in Lithuanian groundwater ranges from approximately 1 µg/L to nearly 39 µg/L with median equal 7.0 (IQR 3.5–20) µg/L. Lithium is a natural trace element that is mobilized by rain from rock and soil and dissolves in the drinking water. Compared with drinking water in other regions of the world, the lithium levels observed in Lithuanian and similarly in Danish drinking water are significantly lower and the range is generally more narrow on a European scale [92, 120]. Drinking water derives from groundwater only, and the levels of lithium in groundwater and drinking water are likely stable over time due to the chemical properties of lithium and its geological origin [121, 122]. According to Environmental Protection Agency of Lithuania report, in Lithuania, 82% of population benefits from central water supply networks services. Lithuania relies exclusively on groundwater for drinking water supply [123]. According to the data of the Lithuanian Geological Survey, in Lithuania were registered 1780 wellfields in 2019 [124]. All Lithuanian drinking water is of groundwater origin and clear geographic patterns in lithium levels have been found. There exists only limited data on the temporal variability of lithium in drinking water in Lithuania as well as in other countries. However, based on its chemical properties in aqueous solutions, lithium is considered to be a conservative compound that, due to the Li+ ions’ small size and strong hydration [121], is not expected to change over time or to react chemically with other substances during aeration and sand filtration at the water treatment plants, in the distribution system, or at installations. Recently, a series of ecological studies [22, 24, 25, 27, 28, 30, 93, 98] and meta-analysis [102] has addressed the potential effect of lithium in drinking water on suicide in the general population. Although evidence is pointing in the direction of an inverse association [125], contradictory findings have been reported in different regions and a causal relation is uncertain.

63 Lithium concentration in the natural waters varies and depends on geological, geographic and hydrogeological variables [126]. In some geographic regions, such as the Andes of Northern Argentina, its concentration may reach up to 5.2 mg/L, resulting in daily intake of lithium of up to 10 mg/day [23, 92]. This is relatively high in comparison to the highest level of 1.3 mg/L of lithium measured in Austria [26]. Lithium levels in Texas drinking water ranged between 2.8 and 219 µg/L and were significantly lower than those found in some regions with large natural lithium concentrations [27, 95]. However, they are markedly higher than the levels reported in an East of England study (range: <1–21 µg/L) that found no association [93]. The investigators in that study argued that the narrow range of lithium concentrations could potentially explain their null findings on the association between lithium levels and suicide rates [27]. Although lithium concentrations in ground water varies from <0.05 to 150 μg/L, the reports from the Austria, northern Chile, and northern Argen- tina have showed very high lithium concentrations in drinking water (over 1,000 μg/L) [24, 27, 137]. Other research shows that in temperate humid zones the lithium levels in ground water are lower than in dry, hot regions (about 1,500 μg/L) [138]. There are not drinking water standards for lithium in the European Union. Although, the median value of lithium was evaluated in European bottled waters (9.94 μg/L), we could not find the data of lithium levels in drinking water of the other Baltic or neighbouring countries [139].

4.2. The association between suicide SMR and lithium levels in drinking water and across municipalities

4.2.1. Association between suicide SMR and lithium levels in drinking water: Study I (Pilot study) This is the first study in Eastern Europe investigating the association between suicide rates and lithium levels in public drinking water. The findings from the study carried out for nine Lithuanian municipalities provide confirmatory evidence that higher lithium levels in the public drinking water are nonlinear associated with lower suicide SMR. A significantly inverse association of lithium levels (log transformed) in drinking water were revealed with suicide SMR in the total population and in men. The observed concentrations of lithium ranged from 0.5 to 35.5 μg/L. Our study results indicate that suicide risk varied considerably across the 9 municipalities of Lithuania, based on multivariate modeling that was considering local population size, and comprehensive data on women/men

64 proportion. Our results of the pilot study were consistent with previous studies in that lithium in drinking water was negatively associated with suicide in men [27, 28, 30, 95, 99]. Suicide SMRs were a significantly inverse associated with lithium levels (log transformed) in drinking water for the total suicide SMR (β = −0.28, P = 0.034) and for men (β = −0.70, P = 0.013), but not for women (β = 0.15, P = 0.70) using weighted least squares regression analysis adjusted for the local population size. Lithium levels in the drinking water were variable and ranged from 0.48 to 35.53 μg/L. For example, a large number of samples of drinking water were examined in relation to suicide (SMRs) in about 300 municipalities in Japan [30]. The authors found that lithium levels in drinking water were significantly and inversely associated with men suicide SMRs but not total or women SMRs. This implies that suicide prevention strategies and programs must reflect gender dissimilarities and local circumstances in risk patterns by providing specific localized health intervention strategies, such as reducing suicide risk factors and improving protective factors related to mental health and wellbeing. The natural lithium levels in drinking water have correlated inversely with suicide risk in most [24, 28, 29, 92, 95], but not in all studies of the relationships between lithium levels in drinking water and suicide rates [93, 98]. In Lithuania, men suicide rate is 6 times higher than women [127]. This is consistent with recently published meta-analyses indicating that the female gender is associated with suicide attempts, while the male gender is associated with suicide deaths [46, 128]. Probably, men are more likely to die by suicide because they choose more violent means of death [8, 99, 129, 130]. These gender differences may be explained by the fact that natural lithium intake may influence impulsiveness, a factor that contributes to suicidal behavior [37, 131, 132].

4.2.2. Association between suicide SMR and lithium levels in drinking water: Study II In our previous pilot study [133] we examined a possible relationship between suicide rates and lithium levels in drinking water from 9 cities in Lithuania. We did not detect a significant linear correlation between lithium level and suicide SMR. The onset of the suicide protective action of lithium appears to require at least 15 µg/L level in drinking water. In the study II we included more data on lithium levels from 53 municipalities of Lithuania [133]. Also, we included more variables to examine an association of lithium levels in

65 drinking water and suicide SMRs (total, men, and women). Relationships between lithium levels in drinking water and suicide rates were investigated, adjusting for unemployment rate, visits to psychiatrists, divorce rate, and women/men proportion. An inverted U-Shaped curvilinear relationship was confirmed between higher lithium level in drinking water and lower suicide SMR. Lithium levels in drinking water were significantly negatively associated with suicide SMR in the total population only after it reached the certain level (about 14.5 µg/L). A similar effect of lithium in drinking water was found on women suicide SMR, with no such correlation observed for men. We found that lithium level in drinking water was significantly negatively associated with women suicide SMR only after it reached the level of 13.2 µg/L. A similar effect of lithium in drinking water on men suicide SMR was marginally significant. Our finding is consistent with several previous studies, which indicated that there are gender differences in the effects of lithium in drinking water on suicide rates [24, 27, 28, 98]. These gender differences may be explained by the fact that natural lithium intake may influence impulsivity, a factor that contributes to suicidal behavior [37, 131, 132]. Another factor influencing higher suicide rates among men is the fact that they tend to be more affected by unemployment or divorce [134]. Lithium is a substance that seems to possess neuroprotective abilities by modulating a large array of intracellular cascades and pathways involved in oxidative stress, inflammation, mitochondrial dysfunction, membrane ho- meostasis, and inhibitory effects on glycogen synthase kinase 3 [33, 72, 117, 135, 136]. Lithium induced changes in signalling pathways regulate changes in protein and gene expression, resulting in long-term effects [137, 138]. Although the effects of therapeutic doses of lithium are well established, little is known about the health effects of natural lithium intake. Lithium occurs naturally in drinking water worldwide in much lower doses, with large geographical variation. Across Europe, the majority of public tap water is derived from groundwater, with some countries including Denmark, Austria and Lithuania supplying almost 100% groundwater to their consumers [139]. Further, long-term intake of micro doses of lithium such as those found in drinking water, in the range 5–50 μg/L, have been found to correlate inversely with suicide risk in most [24, 28, 29, 92, 95] but not in all studies of the relationships between lithium levels in drinking water and suicide rates [93, 98]. Previous studies of the association between lithium concentration in drinking water and suicide rates were criticized for using inadequate statistical methods and neglecting socio-economic confounders [140]. A nationwide Austrian study (lithium levels range: <1–130 µg/L), found an inverse association between lithium levels in drinking water and suicide rates after

66 adjustment for population density, per capita income, proportion of Roman Catholics, as well as the availability of mental health service providers [26]. Recently, an Austrian group reconfirmed an inverse association between lithium levels in drinking water and suicide rates after adjustment for county- based population density, age, gender, race/ethnicity, median income per household, poverty and unemployment rates in Texas [27]. Lithium levels in Texan drinking water ranged between 2.8 and 219 µg/L and were significantly lower than those found in some regions with high natural lithium concentrations [27, 95]. However, they are markedly higher than the levels reported in an East of England study (range: <1–21 µg/L) that found no association [93]. The investigators in that study argued that the narrow range of lithium concentrations could potentially explain their null findings on the association between lithium levels and suicide rates [27]. The results of our study are somewhat consistent with findings by Knudsen et al. (2017) [97]. This study in Denmark is the first to investigate the effect of naturally occurring lithium in drinking water on the incidence of suicide at an individual level with 22 years of follow-up [97]. The authors found that there does not seem to be a protective effect of exposure to lithium in drinking water on the incidence of suicide with levels below 31 μg/L. Similarly, our study shows that an anti-suicidal effect of lithium in drinking water is not present if the lithium concentration is below a certain level. A substantial amount of data shows that lithium significantly reduces mortality by suicide with evidence of efficacy for both short-term and long- term treatment [17]. The amount of lithium found in drinking water is much lower than the dose of lithium salts considered therapeutic in psychiatric practice. In a nation-wide population-based study, Kessing et al. (2017) investigated whether long-term intake of micro levels of lithium in drinking water correlates with the incidence of bipolar disorder [141], and with the incidence of dementia [142] in the general population, hypothesizing an inverse association in which higher long-term lithium exposure is associated with lower incidences of bipolar disorder and dementia. The authors found that higher long-term lithium exposure from drinking water was not associated with a lower incidence of bipolar disorder and they concluded that this association should be investigated in areas with higher lithium levels than in Denmark. However, in another study Kessing et al. (2017) [142] observed that a long-term increased lithium intake with drinking water might be associated with a lower incidence of dementia in a nonlinear way.

67 4.3. The impact of incidence of affective disorders on an association of suicide SMR with lithium level in drinking water

This is the first study to investigate the relationship between suicide SMR, lithium levels in drinking water, socio-demographic characteristics and incidence of MBDs in municipalities simultaneously. Our study suggests a higher incidence of affective disorders in municipalities above median of lithium level in drinking water compared with other municipalities with the same socio-demographic and psychiatric characteristics. In municipalities with above median of lithium level, suicide SMR for total population and among men and women was inversely associated with lithium level in drinking water. The presence of affective disorders could be an important condition for the protective effect of lithium in drinking water against suicide mortality. We found that incidence of affective disorders was higher in municipalities with high lithium exposure (lithium level above median >7 µg/L). The relative difference of incidence of affective disorders was 26.9% and the relative difference of antidepressants use was 19.2% between the low and high lithium exposure groups. In addition, suicide SMR (total, men and women) were negatively associated with lithium level in drinking water in municipalities with high lithium exposure (above the median). Psychiatric inpatients should always be followed-up soon after discharge as periods following hospitalization are often marked with high suicide risk, especially when patients face daily difficulties and cannot rely on family members or any social contact for support [143, 144]. We have theorized that it is likely that if individuals with affective disorders are discharged without lithium treatment in the general population, lithium in drinking water could potentially reduce the rate of suicide among the general population. Lithium treatment remains the “gold standard” of treatment for preventing recurrences in bipolar disorder. There is also evidence of effectiveness for preventing suicidal behavior in patients with bipolar or major depressive disorder. However, it has gradually become less widely utilized, particularly for mania, mainly due to more vigorously promoted and more rapidly effective alternatives, which do not require blood tests. Concerns about the safety of lithium have not entirely disappeared, despite long established standards for its safe use with monitoring of its serum concentrations [145]. Over the last decades, other substances such as second generation antipsychotics (SGA) and anticonvulsants have been prescribed more frequently and there has been a tendency to avoid lithium in the treatment of BD. Reasons may be the overestimation of potential side effects as compared to other substances by professionals and patients alike [145, 146]. The

68 likelihood of patients seeking psychiatric help appears to be related to stigma, still much felt in the Baltic countries, and fear of side effects, over-featured in the variety of Internet sources particularly with regard to antidepressant use [147]. So far, no studies have been found that focused on an association of lithium in drinking water with the incidence of affective disorders in the study region. Our study revealed higher incidence of affective disorder in a sample of III–IV quartiles of lithium level than in lower I–II quartiles with the same suicide SMR and other characteristics of the district municipalities. This suggests that long term exposure to lithium in drinking water has demonstrated possibly specific anti-suicidal effects in those with affective disorders resulting in survival. In addition, multiple regression analysis revealed strong negative association between suicide SMR (total, women and men) and lithium level over median (7 µg/L) controlling for district municipality characteristics and incidence of other mental and behavioral disorders as possible “dual diagnosis”. Unlike most international studies regarding natural lithium levels and suicide risk, no inverse relation was found in Portugal [111] and in Miyazaki Prefecture (Japan) [66]. In these studies, potential confounding factors were socio-demographic data. Factors such as the country's low suicide rate, confounding suicide risk variables, psychiatric disorder, and unaccounted lithium intake might have influenced these findings. The Lithuanian population is ethnically and socially homogeneous and has a very low migration rate. A proportion of patients with affective disorder do not contact a doctor, a proportion may have been treated with poor response to lithium prior to the study period, and a proportion may have been treated with mood stabilizers other than lithium. We are not aware of other national prescribed lithium prevalence rates besides that in Germany, where lithium is estimated to have be prescribed for 0.06% of the German population [73, 148]. According to the recent review of Sarai and coworkers (2018) [149] the anti-suicidal properties of lithium might be unrelated to the mood-stabilizing effects. Affective disorders are a common psychopathology and are a significant risk factor for suicidality. Lithium pharmacotherapy has been shown to reduce symptoms of suicidal behavior, especially in long-term patient interventions. Reasons for this remain unclear. Lithium treatment for individuals with affective disorders appears underutilized. Use of lithium is thought to reduce risk for suicidality, even if mood stabilization is not achieved and serum concentration is lower than the conventionally accepted therapeutic blood level ranges. This brief report serves as an important

69 reminder to clinicians to include lithium pharmacotherapy in their armamentarium for treatment of affective disorders, especially when symptoms of suicidality are present. The anti-suicidal aspects of lithium appear to be as effective at low concentrations as at therapeutic levels [150]. Sustained low doses of lithium intake have been shown to decrease suicidality [128]. There is a significant reduction in the number of suicide attempts among individuals who take lithium, even in those who do not respond well to lithium’s mood symptom prophylaxis [150, 151]. A relationship between lithium levels in tap water and a reduced suicide risk has been postulated multiple times, which a more recent study did not replicate [125]. The mechanism by which lithium may reduce suicides is hypothesized to be the reduction of impulsive and aggressive behavior in bipolar and depressed patients. Some authors suggest, it is thought to have a specific anti-suicide effect exceeding its mood stabilizing properties [146, 152]. On the other hand, although lithium levels are extremely low in drinking water, long-term exposure to lithium may be a factor which mitigates low absolute levels. It can be speculated that very low but very long lithium exposure can enhance neurotrophic factors, neuroprotective factors and/or neurogenesis, which may account for a reduced risk of suicide [153]. Some researchers have even theorized that adding lithium to drinking water could potentially reduce the rate of suicide among the general population [150, 151]. However, the actual concentration needed to induce anti- suicidal effects in people with affective disorders remains unclear [154]. The findings of our study also raise the interesting possibility that it is likely that in the absence of prescribed lithium in the population for the treatment of affective disorder, lithium in drinking water may have an anti- suicidal effect for those with affective disorder. The study by Lewitzka et al (2015) also suggests that the expected higher overall mortality in patients with affective disorders using lithium is decreased [151]. Our data of incidence of affective disorders were collected from the Health Information Centre of the Institute of Hygiene (Lithuania Database of Health Indicators). This means that all cases have been diagnosed and, given that those at high risk of suicide are usually hospitalized and have been registered, lithium has demonstrated possible specific anti-suicidal effects apart from its prophylactic efficacy: it significantly reduces the high excess mortality of patients with affective disorders. The physiological effects precipitated by lithium exposure that tend to reduce suicidal behavior remain unclarified. It is important to learn more about the level of lithium required for it. The literature supports the efficacy of long-term lithium treatment for suicidal patients [19, 155, 156], and lithium

70 is considered to be the first-line treatment for reducing the risk of suicide [157] while there is weaker evidence for the usefulness of anticonvulsants in preventing suicide [155, 157]. There is strong evidence that suicide mostly occurs among people with affective disorders [7]. The suicide risk has been estimated at 6–10% in the mood disorder population, which is 10 times the corresponding risk in non- psychiatric populations [8]. The brief report of Sarai (2018) serves as an important reminder to clinicians to include lithium pharmacotherapy in their armamentarium for treatment of affective disorders, especially when symptoms of suicidality are present [100, 149]. Unemployment In our study, the multiple regression analysis revealed that the total suicide SMR was negatively associated with lithium in drinking water, and a significant positive relationship was found with the proportion of unemployment. A meta-analysis has shown that unemployment impairs mental health. However, 64% of people with common mental health problems are employed [158]. Being unemployed was associated with a twofold to threefold increased relative risk of death by suicide, compared with being employed. Unemployment has been associated with an increased likelihood of suicide, with this association being greater for men than for women [159]. About half of this association might be attributable to confounding by mental illness [160]. The negative effect of unemployment on mental health has been found to be stronger in countries with more unequal income distributions (controlling for level of economic development) [158, 161]. As in other studies, Agerbo and colleagues found that suicide rates increased with increasing income among patients [162, 163]. This might be the effect of increased stigma [164] or because employed patients are in a particularly stressful situation. There are ongoing questions about whether unemployment has causal effects on suicide as this relationship may be confounded by past experiences of mental illness (e.g. becoming unemployed as a suicidal crisis). Results of the meta-analysis of Milner et al (2014) showed that unemployment was associated with a significantly higher relative risk (RR) of suicide before adjustment for prior mental health [RR 1.58, 95% CI 1.33–1.83]. After controlling for mental health, the RR of suicide following unemployment was reduced by approximately 37% [RR 1.15, 95% CI 1.00–1.30]. Greater exposure to unemployment was associated with higher RR of suicide, and the pooled RR was higher for men than for women. This review quantified the effects of adjustment for mental health on the relationship between unemployment and suicide [161].

71 The findings support the idea that unemployment or lack of job security increases the risk of suicide and that social and economic policies that reduce unemployment will also reduce the rate of suicide. The risks in recommending reduced unemployment as a remedy for suicide are twofold: inefficiency (lacking the causal link, one might invest large resources without reducing suicide rates) and lack of focus (by paying attention to phenomena only marginally connected with the problem, we might lose the focus from more credible forms of prevention) [165]. Gender differences Suicide victims have been found to frequently suffer from mental disorders, often more than one, and comorbidity has also been found to be a risk factor for suicide. There are different factors associated with suicide depending on gender, which may also point to different psychological mechanisms in the background of suicide. In our study the highest local R2 values are found in the SMR women, which explained approximately 42% of the variation in the suicide SMR. In the case of women lithium and disease of the nervous system (negatively) and schizophrenia (positively) is more strongly associated with suicide mortality rate. In our study MBD due to use of alcohol in women was the most important factor suicide SMR for men. Suicide and alcohol use Alcohol abuse were associated with diagnoses of several types of personality disorder and bipolar disorder and presented a greater suicide risk than the subgroup of other MBDs due to use of alcohol [166]. MBD due to use of alcohol was the most common first mental disorder among male suicide victims and could thus be considered a starting point in the suicidal process. In addition to detecting and treating depression, it is important to detect and treat MBD due to use of alcohol vigorously and to be alert for subsequent symptoms of depressive and other mental disorders in suicide prevention efforts [167]. Bipolar disorder has been reported to have the greatest risk of any Axis I disorder for coexistence of an alcohol or drug use disorder [168, 169]. Those with a substance abuse/dependence history were more likely to be male, divorced, separated or widowed. Those with bipolar affective disorder and substance abuse are more likely to have mixed or dysphoric manic states [169]. One question, given the high risk for suicidal behavior in mixed states, is the role of prophylactic lithium in patients susceptible to mixed states. Notably data show that lithium appears to reduce suicidal behavior even when it is not effective in preventing affective episodes [170]. A condition when alcohol dependence is accompanied by another mental disorder is much more prevalent believed. It is estimated that more than one

72 third of people diagnosed with mental disorders, abuses or is dependent on psychoactive substances, especially alcohol; among alcohol-dependent patients 37% suffer from other mental disorders. Alcohol dependence is associated with increased risk of mood disorders – more than 3 times higher, depression – almost 4 times higher, bipolar disorder – more than 6 times higher, anxiety disorders in general – more than twice, generalized anxiety disorder – more than 4 times higher, panic disorders – almost double, posttraumatic stress disorder – more than twice. Underestimation of comorbidity is an important problem during treatment of this population of patients. Social skills training can improve stress management and decrease alcohol and drug use among dual diagnosed patients. Researchers studying dual diagnosis underline the fact that simulta- neous treatment of alcohol dependence and co-occurring psychiatric disorders increases the chance to improve patients` functions. Inappropriate treatment without complete management of all existing problems may make full recovery impossible [171]. Mental and behavior disorders Understanding, prediction and prevention of suicidal behavior is one of the most challenging tasks in society in general and in psychiatry. It has become a priority in particular during recent years, as several psychological autopsy studies of suicide victims have shown that the majority were suffering from a mood disorder, usually major depression, with frequent comorbidity of various other mental disorders (in particular anxiety disorders) [74, 172, 173]. The relationship between suicide and MBDs is an important issue. More than two-thirds of suicide completers and suicide attempters have had mostly untreated major depressive episodes at the time of the suicidal act [96]. Del Matto et al. (2020) conducted a systematic review aimed at summa- rizing evidence on the use of lithium for the prevention of suicide risk both in affective disorders and in the general population. Most of the observational studies reported a reduction in suicide in patients with mood disorders. All studies about lithium treatment's duration reported that long-term lithium gave more benefits than short-term lithium in suicide risk. The evidence seems to attribute an intrinsic anti-suicidal property to lithium, independent of its proven efficacy as a mood stabilizer [174]. In our samples, however, we had no access to medical records regarding past pharmacotherapy and history of prescribed lithium, therefore cannot know if any causal relationship exists between the variables. This could be important because there is growing evidence for suicidal risk reduction with long-term lithium maintenance [19, 155, 156, 175]. The relationship between suicide and MBDs is an important issue [143, 144, 176].

73 However, as Hendin (1986) pointed out: “the vast majority of depressed, schizophrenic, alcoholic or organically psychotic patients do not commit or even attempt suicide”. Hendin went on to suggest that “the interest in classifying populations of suicidal patients by their psychiatric diagnoses is being supplemented by an interest in understanding what makes a minority of patients within any given diagnostic category suicidal while the majority are not suicidal” [177, 178].

74 LIMITATIONS AND STRENGTH OF THE STUDY

Limitations. Firstly, the main limitation of the present study is due to the nature of observational research which cannot reveal a causal relationship. Secondly, the limitations of our study include the lack of data relevant to lithium levels in food and the proportion of the population who drank tap water and their consumption habits. We do not know/cannot take into account people’s daily job/living area migration patterns or their tap water consumption tendencies while in different areas. On the other hand, lithium rich food often comes from worldwide market, while drinking water usually has a local origin. Thirdly, other factors such as psychosocial and economic factors were not taken into consideration. We could not obtain data on how many suicides committed people actually had lithium treatment and how long it took. Thus, the findings for association of lithium in drinking water and suicide mortality rate could be biased by possible lithium treatment, if any, and therefore not solely affected by natural lithium occurrence. It should be noted that ecological studies per se are designed to establish hypotheses rather than to provide cause, and their results are not applicable to individual cases (ecological fallacy). Thus, although informative, the estimates should be interpreted with caution due to the aggregated nature of data [26]. Finally, the use of the term “nonlinear” in this study refers to the relationship, not the statistical model. We discussed how such relationships can be better theorized and tested. The examined model of suicide mortality is subject for further interesting research questions. Since all data were not collected in a prospective fashion, the models developed can only be used to test for an association between these independent variables and the outcome; rather than to identify and determine the risk factors or determinants for suicide SMR. Official data on the registered incidence of mental and behavioral disorders and suicide rates are also relatively uninformative due to their incompleteness and the lack of specific epidemiological surveys [52]. Limitations include the use of cross-sectional data. The present study was based on the correlations among suicide SMR, a relatively limited number of discrete diagnoses of mental and behavioral disorders, and the central public drinking water wellfields of municipalities.

75 Strengths. We report here, for the first time, the existence of an inverted U-shape relationship between lithium levels in drinking water and suicide mortality rates. The examined model of suicide mortality is an example for further interesting research questions. No recent studies have been found that analyse the association of lithium level in drinking water with the incidence of mental and behavioral disorders in the study region. This is the first report to investigate the relationship between suicide SMRs, lithium levels in drinking water, socio-demographic characteristics and incidence of affective disorders in municipalities simultaneously.

76 CONCLUSIONS

1. The mean lithium concentration in the public drinking water samples of central wellfields in Lithuanian municipalities was 11.5 (SD 9.9) µg/L ranging from 1.0 to 39.0 µg/L, median – 7.0 (IQR 3.5–20) µg/L. Lithium levels in central drinking water systems are differently distributed across Lithuania. The lowest lithium levels are observed in the Eastern Lithua- nia, Žemaitija and Šilutė, while the highest in regions of Central Lithua- nia, Northern Lithuania and Klaipėda.

2. Lithium levels in the public drinking water are associated with suicide SMR in a nonlinear way. The inverted U-shaped curvilinear relationship is confirmed between higher lithium level in drinking water and lower suicide SMR even after controlling for socio-demographic characteristics. Lithium levels in drinking water are negatively associated with suicide SMR only when the lithium level is higher certain value (14.5 µg/L). A similar effect of lithium in drinking water was is found on women suicide SMR, with no such association observed for men.

3. Lithium levels in drinking water are positively associated with the incidence of affective disorders. The present findings suggest higher incidence rates of affective disorders in the municipalities with a lithium level in drinking water above median compared to the municipalities with a lithium level below median and with the same socio-demographic and psychiatric characteristics. Suicide SMRs are inversely associated with lithium levels in drinking water only in municipalities with higher lithium levels (above median) and with a high rate of affective disorders.

Based on our study results and insights we generated the following hypothesis for the further research, that lithium level in drinking water might have an important protective effect to against suicide mortality in the population with affective disorders.

77 PRACTICAL AND SCIENTIFIC RECOMMENDATIONS

The increasing problem of suicides and mental disorders in developed countries creates an urgent need to develop novel prophylactic strategies to protect mental health. According to our study results, and other numerous observations, the intake of lithium from drinking water, may negatively affect suicide rates, however this hypothesis requires further in-depth research investigating the mechanisms of action of trace doses of lithium. We do not claim here any causality, but we do hope that the findings of the study can be of help to policy makers in reducing the incidence of suicide in the country in the future. Practical. Our study results highlight the need for mental health policy makers to be informed about possibly specific anti-suicidal effects of lithium levels in drinking water, emphasizing the relationship with the incidence of affective disorders and suicide rates. It is necessary for health policy makers to pay attention to regional inequalities in suicide rates when implementing suicide prevention measures for people with mental health disorders, focusing on the municipalities where the highest number of suicides remains. Scientific. It is still not clear what amount of lithium in drinking water can provide an independent protective effect against suicide. In particular, more research is needed to understand how lithium levels in the public drinking water supply correlate with blood lithium levels. More research would help draw conclusions about trace lithium’s anti- suicidal effect, and provide new knowledge for future research studies evaluating biological mechanisms on suicidal behavior at the population level (among people with affective disorders). It should be focus to individuals with known increased suicide risk, such as patients diagnosed with psychiatric disorders (affective disorders) or with other medical conditions linked to higher risk of suicide. Also, should be evaluate the lithium concentration in drinking water, examining not only central but also less commonly used watering-place and individual wells, especially in municipalities with high rates of suicide (affective disorders). Comprehensive studies should be initiated for evaluating the links between lithium and diseases of nervous systems, and/or other health- related indicators.

78 The knowledge of the beneficial properties of lithium and its role on suicidal behavior can lead to a deeper understanding of mental illness and improve the wellbeing of patients with affective disorders and other mental health problems. Despite the need for future research, this study adds further in-sight on the relationship between natural lithium levels in drinking water and suicide mortality.

79 SANTRAUKA

1. Įvadas

Savižudybė yra svarbi visuomenės psichikos sveikatos problema Lietuvoje ir visame pasaulyje [55]. Savižudybė yra antra pagal 15–29 metų amžiaus gyventojų mirties priežastis visame pasaulyje, kasmet dėl šios priežasties pasaulis netenka maždaug 800 000 gyvybių [55]. Savižudybė yra sudėtingas reiškinys, apimantis daugybę psichologinių, socialinių, ekonomi- nių, biologinių ir aplinkos veiksnių [136]. Tyrimai rodo, kad psichikos sutrikimai gali padidinti savižudybių riziką nuo 3 iki 12 kartų [3], tuo tarpu nuotaikos (afektiniai) sutrikimai labiausiai susiję su savižudišku elgesiu [6, 110]. Nustatyta, kad savižudybės rizika afektinių sutrikimų populiacijoje siekia 6–10 proc. ir yra 10 kartų didesnė negu bendrojoje populiacijoje [55]. Pacientų su savižudiško elgesio rizika gydymas yra viena iš sunkiausių užduočių, tenkančių sveikatos priežiūros specialistams. Iki šiol buvo siūlo- mos kelios savižudybių prevencijos strategijos populiacijos ir individualiu lygmeniu, kai kurios iš jų – farmakologinės [55]. Klinikiniais tyrimais įrody- ta, kad litis yra vienas efektyviausių vaistų depresijos ir bipolinio sutrikimo gydymui, bei suicidinio elgesio prevencijai [148]. Sisteminė atsitiktinių imčių tyrimų apžvalga pateikė išvadas, kad gydymas ličiu sumažina mirtingumą nuo savižudybės daugiau nei 60 proc. sergančiųjų sunkia depresija ar bipoliniais sutrikimais [55]. Placebu kontroliuojamų tyrimų duomenys parodė, kad mažos ličio dozės gali stabilizuoti nuotaiką pacientams, turintiems priklausomybę nuo psichotropinių medžiagų [14]. Savižudybių prevencijai svarbi ir gydymo ličiu trukmė. Klinikiniai atsitiktinių imčių tyrimai pateikia išvadas, kad ilgalaikis gydymas ličiu prailgina pacientų, sergančių afektiniais sutrikimais, išgyvena- mumą, bendrojoje populiacijoje. Pacientų, sergančių bipoliniais ir afektiniais sutrikimais, gydymui naudojant litį vidutiniškai 18 mėnesių, suicidinių bandymų ir mirčių nuo savižudybės rizika sumažėja apie 80 proc. [67]. Nors aplinkoje randama ličio koncentracija yra daug kartų mažesnė už terapinę ličio dozę psichikos sutrikimų gydymui [67], vis daugiau pateikiama įrodymų, kad net labai maža ličio koncentracija geriamajame vandenyje gali turėti antisuicidinį poveikį tiek pacientams turintiems afektinių sutrikimų, tiek bendrajai populiacijai [142]. Viena iš hipotezių, aiškinančių mažos ličio koncentracijos antisuicidinį poveikį, yra ta, kad būtent ilgalaikis ličio vartojimas (su geriamuoju vandeniu) derinyje su maža ličio koncentracija gali paaiškinti mažos ličio koncentracijos antisuicidinį poveikį [142].

80 Epidemiologiniai tyrimai pateikia išvadas, kad geriamajame vandenyje esanti ličio koncentracija gali būti susijusi mažesniais mirtingumo nuo savižudybių rodikliais [24–31], mažesniais demencijos sergamumo rodikliais, mažesne depresijos rizika bei agresijos raiška paaugliams [32, 33]. Ličio antisuicidinis poveikis gali išryškėti dėl jo nuotaiką stabilizuojančių savybių, mažinant agresyvumą ir impulsyvumą, kurie siejami su padidėjusia savižu- dybės rizika [148]. Nepaisant šio susidomėjimo, nei viename tyrime, kiek mums žinoma, netirta sąsaja tarp psichikos ir elgesio sutrikimų paplitimo, ypač tarp sergamumo afektiniais sutrikimais ir ličio antisuicidinio poveikio geriamajame vandenyje tiriamos populiacijos lygmenyje. Iškėlėme hipotezę, kad didesnė ličio koncentracija geriamajame vandenyje turi apsauginį poveikį mirtingumui nuo savižudybių savivaldybėse, kuriose didelis sergamumas afektiniais sutrikimais.

2. Tyrimo tikslas ir uždaviniai

Darbo tikslas – ištirti ličio koncentraciją geriamajame vandenyje, ir nustatyti jos sąsajas su savižudybių ir afektinių sutrikimų rodikliais.

Darbo uždaviniai: 1. Ištirti ličio koncentraciją geriamajame vandenyje Lietuvos savival- dybių centrinėse vandenvietėse. 2. Nustatyti mirtingumo nuo savižudybių sąsają su ličio koncentracija geriamajame vandenyje. 3. Įvertinti afektinių sutrikimų paplitimo sąsają su ličio koncentracija geriamajame vandenyje.

Ginami teiginiai 1. Ličio koncentracija geriamajame vandenyje skiriasi tarp Lietuvos savivaldybių vandenviečių. 2. Didesnė ličio koncentracija geriamajame vandenyje susijusi su mažesniais mirtingumo nuo savižudybių rodikliais. 3. Ličio koncentracija geriamajame vandenyje susijusi su mirtingumo nuo savižudybių rodikliais savivaldybėse, kuriuose didesnis afektinių sutrikimų paplitimas.

81 Darbo mokslinis naujumas Tai pirmas ekologinis tyrimas Rytų Europoje, vertinantis ličio koncentracijos geriamajame vandenyje ir mirtingumo nuo savižudybių sąsają. Šiame tyrime buvo tikrinama hipotezė, kad didesnė ličio koncentracija geriamajame vandenyje yra susijusi su mažesniu mirtingumu nuo savižudybių tarp šalies savivaldybių, kuriose yra didelis sergamumas afektiniais sutrikimais. Antra, šiame tyrime pirmą kartą nustatytas apverstos U formos ryšys tarp ličio koncentracijos geriamajame vandenyje ir mirtingumo nuo savižudybių rodiklių populiacijos (šalies savivaldybių) lygmeniu. Nagrinėjamas mirtingumo nuo savižudybių rodiklių modelis yra tolimesnių naujų tyrimų subjektas. Trečia, iki šiol, mūsų žiniomis, nebuvo atlikta tyrimų, kuriuose ličio koncentracijos geriamajame vandenyje sąsaja su mirtingumu nuo savižudy- bių būtų siejama su sergamumu afektiniais sutrikimais, keliančiais didelę savižudybės grėsmę. Šiuo tyrimu, pirmą kartą buvo vertinama sąsaja tarp mirtingumo nuo savižudybių ir ličio koncentracijos geriamajame vandenyje, kontroliuojant sociodemografinius veiksnius ir sergamumo psichikos ir elgesio bei kitais sutrikimais turinčiais galimą savižudybės riziką rodiklius savivaldybių lygmeniu. Mūsų tyrimas atskleidė didesnį sergamumą afektiniais sutrikimais savivaldybėse, kuriose ličio koncentracija geriamajame vandenyje viršijo visų ištirtų mėginių ličio koncentracijos medianą (mūsų tyrime ≥7 µg/L). Todėl atrodo tikėtina, kad ličio poveikis yra ne tik nuotaiką stabilizuojantis, bet ir turintis antisuicidinį poveikį esant ir labai mažai ličio koncentracijai (lyginant su terapinėmis ličio dozėmis). Nors geriamajame vandenyje ličio koncentracija yra labai maža, ilgalaikis ir pastovus ličio pasisavinimas su geriamuoju vandeniu, gali būti veiksnys, padidinantis mažos ličio koncentracijos poveikį. Kita vertus, šis tyrimas pateikia svarbią informaciją apie ličio koncentracijos geriamajame vandenyje ir SMR nuo savižudybių sąsajas. Tai gali suteikti naujų įžvalgų būsimiems tyrimams, kuriuose būtų vertinami sergan- čių afektiniais sutrikimais suicidinio elgesio biologiniai mechanizmai. Galiausiai, tyrimas patvirtina, kad savižudybė yra labai daugiasluoksnis fenomenas, kuriam paaiškinti neužtenka vieno modelio, o ši disertacija prisideda prie gilesnio jo pažinimo per menkai tyrinėtas sąsajas.

Autoriaus asmeninis indėlis Disertacinio darbo autorė nuo pat tyrimo pradžios aktyviai dalyvavo renkant geriamojo vandens mėginius pirmame tyrimo etape. Tiesiogiai bend- ravo su savivaldybių centrinių geriamojo vandens vandenviečių administra- cijos darbuotojais dėl leidimo rinkti vandens mėginius mikroelemento ličio

82 tyrimui. Tyrimo metu autorė parengė projektą, skirtą visų Lietuvos rajonų savivaldybių centrinių geriamojo vandens vandenviečių ličio koncentracijos ištyrimo finansavimui gauti. Darbo autorė taip pat dalyvavo antrajame tyrimo etape, organizuojant vandens mėginių rinkimą bei ištyrimą su Vilniaus universiteto Gamtos tyrimų centro atstovais, ir laboratorijos atstovais – mėgi- nių ištyrimui. Autorė pati apdorojo Lietuvos savivaldybių gyventojų socialinius- demografinius, sergamumo ir mirtingumo rodiklius, naudodamasi tiesioginės prieigos prie Lietuvos sveikatos rodiklių duomenų baze. Atliko duomenų statistinę analizę ir apibendrino gautus rezultatus. Doktorantūros metu autorė pristatė darbo rezultatus tarptautinėse ir nacionalinėse konferencijose Lietuvoje ir užsienyje. Gautus rezultatus paskelbė recenzuojamuose užsienio žurnaluose.

3. Tyrimo medžiaga ir metodika

Tyrimo eiga Atliktas ekologinis momentinis tyrimas, kuriuo analizuojama sąsaja tarp mirtingumo nuo savižudybės ir ličio koncentracijos geriamajame vandenyje, šalies savivaldybių lygmenyje. Tyrimas atliktas dviem etapais pagal pateiktą tyrimo tėkmės schemą (3.1.1 pav.). Tyrimo objektas ir metodika Tyrimo objektas buvo savivaldybės (miesto arba rajono) vandenvietės. Ličio koncentracijos tyrimui geriamojo vandens mėginiai buvo paimti iš kiekvienos rajono arba miesto savivaldybės centrinės vandenvietės, pagal didžiausią išgaunamą (eksploatuojamą) požeminio vandens kiekį kiekvienoje savivaldybėje. Vandens mėginių paėmimo metodika ir vandenviečių atranka buvo aptarta ir suderinta su Gamtos tyrimų centro geologais.

3.1.1 pav. Tyrimo tėkmės schema *II tyrime, 1 vandenvietės geriamojo vandens mėginio rezultatai buvo pašalinti iš statistinės analizės dėl labai didelės ličio koncentracijos (49.0 µg/l Šakių savivaldybės centrinėje vandenvietėje). Šis rodiklis buvo pašalintas iš duomenų bazės dėl išskirties (3 priedas).

Ličio koncentracijos geriamajame vandenyje matavimai I (pilotinis) tyrimas, atliktas 2013 m. lapkričio mėn.–2014 m. sausio mėn. Siekiant kuo labiau sumažinti ekonominės ir kultūrinės aplinkos nevienalytiš- kumo poveikį, geriamojo vandens mėginiai buvo paimti tik iš miestų (ne rajonų) vandenviečių. Šiam tyrimui buvo pasirinkti 5 didieji miestai: Vilnius, Kaunas, Klaipėda, Šiauliai, Alytus (išimtis buvo Panevėžys, nes negavome

84 atsakymo į paklausimą dėl mėginių paėmimo), ir 4 kurortiniai miestai: Palan- ga, Neringa, Birštonas, Druskininkai). Tyrimo metu buvo surinkti ir ištirti 22 geriamojo vandens mėginiai (nuo 1 iki 5 mieste) iš visų (22) vandenviečių esančių miestuose. Ličio koncentracijos tyrimai buvo atlikti Lietuvos sveikatos mokslų universiteto Neuromokslų instituto Toksikologijos laboratorijoje, pagal indukty- viai surištos plazmos masių spektrometrijos metodiką (ICP-MS) NexION™ 300D (PerkinElmer, USA). II tyrimas atliktas 2017 m. birželio–liepos mėn. Tyrimo metu buvo ištirti 56 geriamojo vandens mėginiai (po 1 kiekvienoje savivaldybėje, išskyrus Klaipėdos savivaldybėje – 3) iš 56 centrinių vandenviečių visose šalies savivaldybėse, įskaitant miestų savivaldybes (Druskininkai, Birštonas, Klai- pėda, Neringa, Palanga, Kalvarija, Kazlų Rūda, Marijampolė, Rietavas, Visa- ginas, Elektrėnai) ir rajonų savivaldybes (Alytus, Lazdijai, Varėna, Jonava, Kaišiadorys, Kaunas, Klaipėda, Kėdainiai, Prienai, Raseiniai, Klaipėda, Kretinga, Skuodas, Šilutė, Šakiai, Vilkaviškis, Biržai, Kupiškis, Panevėžys, Pasvalys, Rokiškis, Akmenė, Joniškis, Kelmė, Pakruojis, Radviliškis, Šiauliai, Jurbarkas, Šilalė, Tauragė, Mažeikiai, Plungė, Telšiai, Anykščiai, Ignalina, Molėtai, Utena, Zarasai, Šalčininkai, Širvintai, Švenčionys, Trakai, Ukmergė, Vilnius). Klaipėdos miesto ir rajono savivaldybėse buvo paimti 2 papildomi mėgi- niai dėl skirtingo eksploatuojamo vandeningojo sluoksnio, naudojamo viešo- jo vandens tiekimui. Ličio koncentracijos tyrimai buvo atlikti UAB „Vandens tyrimai“ laboratorijoje, taikant chromatografijos metodiką, naudojant jonų chromatografą DIONEX ICS-1000 (Thermo Scientific, USA). Grafinis ličio koncentracijos reikšmių vaizdavimas stačiakampe diagrama leido nustatyti išskirtį, labiausiai nutolusią nuo medianos (3 priedas). Viename geriamojo vandens mėginyje ličio koncentracija geriamajame vandenyje siekė 49 µg/l (Šakių vandenvietėje). Atmetus šį rodiklį kaip išskirtį, tyrimo imtį sudarė 55 vandenviečių mėginiai 53 savivaldybėse. Visi geriamojo vandens mėginiai ličio koncentracijai nustatyti buvo imami į 50 ml talpos vienkartinius specialius PET buteliukus, skirtus mikro- elementų analizei atlikti. Surinkti mėginiai buvo laikomi šaldytuve, ir pristatyti į laboratorijas dviejų dienų laikotarpyje. Priklausomas kintamasis: mirtingumo nuo savižudybių rodikliai Mirtingumo nuo savižudybių (SMR, standartizuotas mirtingumo rodiklis) rodikliai savivaldybėse per 2009–2013 m. ir 2012–2016 m. laikotarpius buvo surinkti naudojant Higienos instituto Sveikatos statistinių duomenų tiesioginės prieigos portalą.

85 Nepriklausomi kintamieji Savivaldybių (miestų arba rajonų) sociodemografiniai rodikliai (vidutinis gyventojų skaičius, moterų skaičius tenkantis 1000 vyrų; nedarbo lygis (proc.); apsilankymų pas psichiatrus skaičius tenkantis 100 gyventojų; skyry- bų skaičius tenkantis 1000 gyventojų), mirtingumo nuo savižudybių (SMR, standartizuotas mirtingumo rodiklis), sergamumo psichikos ir elgesio sutrikimais, afektiniais sutrikimais, šizofreniniais sutrikimais, psichikos ir elgesio sutrikimais vartojant alkoholį, nervų sistemos ligomis rodikliai, ir bandymų žudytis skaičius bei antidepresantų vartojimo skaičius tenkantis 1000 gyven- tojų per dieną, surinkti naudojant Higienos instituto Sveikatos statistinių duomenų tiesioginės prieigos portalą.

3.3. Statistinė analizė

Statistinė duomenų analizė atlikta SPSS programine įranga (SPSS Inc, Chicago, IL, USA) „SPSS v. 17.0“. Buvo taikyti standartiniai aprašomosios ir analitinės statistinės duomenų analizės metodai: grafinė duomenų analizė, išskirčių analizė, tiriamų ryšių pobūdžio analizė sklaidos diagramomis, tiria- moji faktorinė analizė kintamųjų skaičiaus sumažinimui. Sąsajoms tarp savižudybių standartizuoto mirtingumo rodiklio (SMR) ir ličio koncentracijos geriamajame vandenyje nustatyti taikyta: mažiausių kvadratų svorinės regresijos modelis, įvertinant skirtingus populiacijų dy- džius; nepriklausomo kintamojo ličio logaritminė, kvadratinė, kvadratinės šaknies transformacija. Logaritminė duomenų transformacija padėjo užtikrin- ti klasikinių tiesinės regresijos prielaidų tenkinimą. Grafinis duomenų vizualizavimas pademonstravo savižudybių SMR ir ličio koncentracijos geriamajame vandenyje apverstos U formos ryšį. Kvad- ratinis ličio terminas (X2) buvo įtrauktas į daugialypės tiesinės regresijos modelį. Norint išvengti multikolineariškumo problemų dėl originalaus ličio kintamojo (X2) ir jo kvadratinio termino (X2) įtraukimo į daugialypės tiesinės regresijos analizę, kintamasis litis buvo centruotas (atėmus vidurkį M) ir sudarytas jo kvadratinis terminas: 2 2 Xc = (litis – M) ir Xc = (litis – M) Kvadratinių terminų reikšmė parodo ryšio netiesinį pobūdį. Ženklas reiš- kia netiesiškumo tipą. Teigiamas kvadratinis terminas gali reikšti, kad ryšys yra eksponentinis. Neigiamas kvadratinis terminas rodo, kad didesnėms reikšmėms ryšys tampa neigiamas. Parabolė aprašoma lygtimi Y = –ax2 + bx + c: lūžio taškas apskaičiuojamas pagal formulę x = –b/2a. Hipotezėms tikrinti ir sąsajų statistiniam patikimumui nustatyti pasirink- tas reikšmingumo lygmuo p<0,05.

86 4. Rezultatai

4.1. Ličio koncentracija geriamajame vandenyje, savižudybių SMR, ir sociodemografiniai savivaldybių rodikliai I (pilotinis) tyrimas Ištyrus 22 vandenvietes 9 savivaldybėse ličio koncentracijos vidurkis buvo 10,9 (SN 9,1) µg/l, svyravo nuo 0,48 iki 35,53 µg/l, mediana – 3,6 µg/l. Bendras savižudybių SMR vidurkis 100 tūkst. gyventojų 2009–2013 m. buvo 27 (ribos 16–50); vyrų – 51 (ribos 29–93) ir moterų – 7 (ribos 0–13).

4.1.1 lentelė. Savivaldybių charakteristikos (N = 9) Populiacija Moterų /vyrų Savižudybių SMR Savivaldybės Litis, µg/lb 2013 m. santykisa Bendras Moterų Vyrų Neringa 2719 1013 50,36 13,3 93,6 1,24 Klaipėda 158 541 1205 19,33 5,7 36,3 13,1 Palanga 15 732 1257 19,05 4,9 37,3 21,79 Šiauliai 106 470 1304 26,88 9,2 50,1 28,68 Kaunas 306 888 1269 21,03 9,3 36,1 11,63 Druskininkai 14 128 1267 29,09 7,6 55,6 3,89 Birštonas 2525 1243 29,56 0 64,9 5,05 Vilnius 526 356 1232 15,97 5,9 29,2 5,87 Alytus 57 281 1177 29,7 7,2 56,8 6,94 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); amoterų skai- čius tenkantis 1000 vyrų; bličio koncentracijos vidurkis.

II tyrimas Ištyrus 55 vandenvietes 53 savivaldybėse ličio koncentracijos vidurkis buvo 11,5 (SN 9,9) µg/l, svyravo nuo 1,0 iki 39,0 µg/l, mediana – 7,0 (IQR 3,5–20) µg/l (4.1.1 pav.).

4.1.1 pav. Ličio koncentracija geriamajame vandenyje, centrinėse Lietuvos savivaldybių vandenvietėse Ličio koncentracijos geriamajame vandenyje geografinis pasiskirstymas pateiktas 4.1.2 pav. Išskiriami šeši rajonai, kuriuose ličio koncentracija geriamajame vandenyje ženkliai skiriasi. Didžiausia ličio koncentracija nustatyta Klaipėdos, Šiaurės ir Vidurio Lietuvos rajonuose.

4.1.2 pav. Ličio koncentracijos geriamajame vandenyje pasiskirstymas pagal rajonus 2012–2016 m. vidutinis gyventojų skaičius savivaldybėse buvo 42 102 (SD 50 452) ir svyravo nuo 2 756 iki 316 916 gyventojų. Bendras, vyrų ir moterų savižudybių SMR vidurkis 100 tūkst. gyventojų 2012–2016 m. ir kiti savivaldybių statistiniai rodikliai pateikti 4.1.2 lentelėje.

4.1.2 lentelė. Savivaldybių sociodemografiniai rodikliai (N = 53) Procentilės Vidurkis SN Min Max 25 50 Mediana 75 Savižudybių SMR bendras 39,5 10,9 16,2 65,5 33,5 39,5 45,0 Savižudybių SMR vyrų 79,0 22,2 31,9 137,1 62,9 75,2 92,5 Savižudybių SMR moterų 13,3 5,1 3,8 28,8 8,8 12,8 15,9 Nedarbo lygis, proc. 25,2 16,8 5,5 17,5 8,9 11,5 13,3 Apsilankymų pas 27,5 8,2 7,3 55,1 22,2 27,5 32,5 psichiatrus sk./100 gyv. Skyrybų sk./1000 gyv. 3,2 0,5 2,3 4,7 2,6 3,1 3,4 Moterų/vyrų santykisa 1136 47 972 1261 1112 1133 1159 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skaičius tenkantis 1000 vyrų; SN, standartinis nuokrypis.

89 Savivaldybių lygmeniu, kaip nepriklausomi kintamieji mirtingumui nuo savižudybių, buvo analizuojami sutrikimų, turinčių galimą savižudybės riziką sergamumo rodikliai (4.1.3 lentelė).

4.1.3 lentelė. Savivaldybių sergamumo rodikliai, bandymų žudytis skaičius ir antidepresantų vartojimo skaičius (N = 53) Rodikliai, 100 000 gyv. Min Max Vidurkis SN Afektiniai sutrikimai 149,62 991,47 474,43 226,68 Nervų sistemos ligos 1019,11 8762,92 4424,28 1554,20 Šizofrenija 14,77 110,91 49,65 23,403 Psichikos ir elgesio sutrikimai (PES) 1457,27 4194,89 2893,79 655,72 Bandymų žudytis sk. 13,30 108,90 52,21 24,36 Antidepresantų vartojimas / 1000a 1,00 35,8 19,97 8,30 PES dėl alkoholio vartojimo, bendras 86,23 528,66 286,41 98,35 PES dėl alkoholio vartojimo, moterų 27,74 215,01 116,01 45,31 PES dėl alkoholio vartojimo, vyrų 160,56 901,37 477,35 173,89 PES, psichikos ir elgesio sutrikimai; SN, standartinis nuokrypis; a antidepresantų vartojimo skaičius tenkantis 1000 gyventojų per dieną.

4.2. Savižudybių SMR ir ličio koncentracijos geriamajame vandenyje sąsajos: I (pilotinis) tyrimas

Grafiškai pateikti duomenys demonstruoja nepriklausomo (litis) ir priklausomo (savižudybių SMR) kintamųjų sąsają tiesiškai ar eksponentiškai. Eksponentinis modelis mūsų duomenims tinka daug geriau nei tiesinis 2 (Rkoreguotas = 0,47; p = 0,025) (4.2.1 pav.).

4.2.1 pav. Ličio koncentracijos ryšys su savižudybių SMR Punktyrinė linija rodo tiesinį ryšį, vientisa – eksponentinę ryšio formą.

Nustatyta bendro ir vyrų savižudybių SMR neigiama koreliacija su ličio koncentracija geriamajame vandenyje (ln litis) ir su moterų/vyrų santykiu. Moterų savižudybių SMR nebuvo susiję su šiais kintamaisiais (4.2.1 lentelė).

4.2.1 lentelė. Savižudybių SMR sąsajos su ličio koncentracija, ir moterų/vyrų santykiu Pearson’o koreliacijos koeficientas (r) Savižudybių SMR Bendras Vyrų Moterų Litis, µg/l –0,430 –0,016 –0,448 Ln litis –0,731* –0,728* –0,276 Moterų/vyrų santykisa –0,770* –0,726* -0,488 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skai- čius tenkantis 1000 vyrų;* p<0,05.

91 Svorinės (kontroliuojant pagal gyventojų skaičių) regresijos analizė išryškino bendro savižudybių SMR neigiamą sąsają su ličio koncentracija (β = –0,91, p = 0,001). Kontroliuojant iškraipantį moterų/vyrų santykio poveikį, ryšys su ličio koncentracija išliko statistiškai reikšmingas (β = –0,28, p = 0,034) (4.2.2 lentelė).

4.2.2 lentelė. Savižudybių SMR ir ličio koncentracijos ryšys (svorinės regresijos modeliai) Savižudybių SMR β t reikšmė p Modelis 1 Litis, µg/L 0,560 1,789 0,117 Modelis 2 Ln litis –0,911 –5,841 0,001 Modelis 3 Ln litis –0,283 –2,886 0,034 Moterų/vyrų santykisa –0,713 –7,319 0,001 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skaičius tenkantis 1000 vyrų.

Vyrų savižudybių SMR buvo neigiamai susiję su ličio koncentracija (β = –0,96, p<0,001), tačiau moterims ši sąsaja nebuvo nustatyta (β = 0,15, p = 0,700). Kontroliuojant iškraipantį moterų/1000 vyrų santykio poveikį sąsaja su ličio koncentracija išliko reikšminga tik vyrams (β = –0,70, p = 0,013) (4.2.3 lentelė).

4.2.3 lentelė. Vyrų ir moterų savižudybių SMR ryšys su ličio koncentracija (svorinės regresijos modeliai: parametrų įverčiai, t kriterijaus statistikos ir p reikšmės) Savižudybių SMR Vyrų Moterų β t p β t p Modelis 2 Ln litis –0,965 –9,684 <0,001 0,150 0,402 0,700 Modelis 3 Ln litis –0,702 –3,515 0,013 0,253 0,677 0,523 Moterų/vyrų santykisa –0,296 –1,484 0,188 0,490 1,311 0,238 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skaičius tenkantis 1000 vyrų.

Nepaisant, kad šis žvalgomasis (pilotinis) tyrimas buvo pagrįstas riboto Lietuvos miestų skaičiaus duomenimis, rezultatai rodo, kad ličio koncentracija geriamajame vandenyje gali būti susijusi su mirtingumo nuo savižudy- bės rodiklių pokyčiais.

92 4.3. Savižudybių SMR ir ličio koncentracijos geriamajame vandenyje sąsajos: II tyrimas Taikant parametrinius metodus, buvo naudojamos kelios ličio kintamojo netiesinės transformacijos, tokios kaip logaritminė, kvadratinė šaknis ir kėlimas kvadratu (4.3.2 lentelė).

4.3.2 lentelė. Savižudybių SMR ir ličio koncentracijos koreliacija Pearson’o koreliacijos koeficientas (r) Originalus ir transformuotas Savižudybių SMR ličio kintamasis Bendras Vyrų Moterų Litis, µg/l –0,150 –0,126 –0,153 Ln litis –0,003 –0,210 –0,084 (Litis)2 –0,253 –0,195 –0,229 Kvadratinė šaknis (Litis) –0,080 –0,076 –0,115 Centruotas: (litis – vidurkis) –0,150 –0,126 –0,153 (Litis – vidurkis)2 –0,397* –0,286* –0,326* SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); *p<0,05.

Bendras, vyrų ir moterų savižudybių SMR reikšmingai neigiamai buvo susiję tik su centruota (vidurkio atžvilgiu) ličio reikšme pakelta kvadratu [(litis-vidurkis)2]. Neparametrinės Spearman’o koreliacijos (4.3.3 lentelė) rodo, kad mirtingumo nuo savižudybių rodikliai reikšmingai koreliuoja su nedarbo lygiu, vidutinių gyventojų skaičiumi ir moterų/1000 vyrų santykiu. Šie trys kintamieji buvo potencialūs kandidatai įtakoti savižudybių SMR taikant daugialypės regresijos modelį. Priešingai, apsilankymų pas psichiatrus skaičius koreliavo tik su moterų savižudybių SMR, o skyrybų skaičiaus rodiklis nekoreliavo su savižudybių SMR.

93 4.3.3 lentelė. Savižudybių SMR ir sociodemografinių rodiklių koreliacijos (Spearman’o koreliacijos koeficientai rs) Savižudybių SMR Bendras Vyrų Moterų

rs p rs p rs p Nedarbo lygis, proc. 0,350 0,010 0,288 0,037 0,173 0,215 Apsilankymų pas psichiatrus sk./ 0,215 0,122 0,209 0,132 0,311 0,024 100 gyv. Skyrybų sk./1000 gyv. –0,169 0,240 –0,053 0,704 –0,001 0,992 Moterų/vyrų santykisa –0,384 0,004 –0,368 0,007 –0,217 0,119 Vidutinis gyventojų sk. –0,322 0,019 –0,308 0,025 –0,162 0,245 SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skaičius tenkantis 1000 vyrų.

Savižudybių SMR ir ličio koncentracijos sąsajos Į daugialypę svorinę mažiausių kvadratų regresiją nepriklausomi kintamieji buvo įtraukti tiesine, laipsnine arba logaritmine forma. Kad būtų aiškesnė taikoma regresinė analizė, iš sklaidos diagramos (4.3.1 pav.) matome, kad pasiekus tam tikrą parabolės tašką – lūžio tašką, didėjant ličio reikšmėms savižudybių standartizuoti mirtingumo rodikliai mažėja.

4.3.1 pav. Savižudybių SMR ir ličio koncentracijos ryšys (sklaidos diagrama) Parabolė aprašoma lygtimi y= –ax2+bx+c: lūžio taškas apskaičiuojamas pagal formulę x= –b/2a.

94 4.3.4 lentelė. Savižudybių SMR ir ličio koncentracijos sąsaja (daugialypės svorinės regresijos analizė) Koeficientaia Nestandartizuotas Standartizuotas p B SE β Modelis 1 (Konstantė) 39,48 2,33

Xc [Litis – vidurkis] 0,355 0,192 0,320 0,070 2 Xc [Litis – vidurkis] –0,049 0,017 –0,482 0,007 Modelio: R2 = 0,175, F = 3,9, p = 0,027 Modelis 2 (Konstantė) 59,98 18,43

Xc [Litis – vidurkis] 0,280 0,173 0,252 0,112 2 Xc [Litis – vidurkis] –0,044 0,018 –0,436 0,018 Nedarbo lygis, proc. 0,876 0,353 1,220 0,017 Apsilankymų pas psichiatrus sk./ 0,482 0,173 0,338 0,008 100 gyv. Modelio: R2 = 0,338, F = 6,30, p<0,001 a Svorinė mažiausių kvadratų regresija dėl populiacijų dydžio skirtumų; Modelis 2: kontroliuotas nedarbo lygio ir apsilankymų pas psichiatrus skaičiaus iškraipantis poveikis; SE, standartinė paklaida.

Tiek bendro, tiek vyrų ir moterų regresinėje analizėje centruoto ličio kintamojo pakelto kvadratu koeficientas buvo statistiškai reikšmingas ir neigiamas. 4.3.4 lentelėje pateikiama bendro savižudybių SMR daugialypės regresijos analizė. Vyrų savižudybių SMR ir ličio koncentracijos sąsajos Išsamesnei duomenų analizei buvo taikyti tiesiniai ir kvadratiniai regresijos modeliai, kad būtų galima nustatyti sritis, kurios gali rodyti tiesinius arba apverstus U formos ryšius tarp ličio geriamajame vandenyje ir vyrų savižudybių SMR (4.3.5 lentelė). Atvirkštinis vyrų savižudybės SMR ryšys su ličio koncentracija geriamajame vandenyje ribinio statistinio reikšmin- gumo (p = 0,055) buvo tik pasiekus 15,3 µg/l lygį.

95 4.3.5 lentelė. Vyrų savižudybių SMR ir ličio koncentracijos sąsaja (daugialy- pės svorinės regresijos analizė) Koeficientaia Nestandartizuoti Standartizuoti p B SE β Modelis 1 (Konstantė) 76,6 4,8

Xc [Litis – vidurkis] 0,702 0,173 0,252 0,086 2 Xc [Litis – vidurkis] –0,075 0,036 –0,365 0,046 Modelio: R2 = 0,083, F = 2,2, p = 0,115 Modelis 2 (Konstantė) 124,8 38,5 0,002

Xc [Litis – vidurkis] 0,554 0,351 0,250 0,118 2 Xc [Litis – vidurkis] –0,073 0,037 –0,355 0,055 Nedarbo lygis, proc. 2,009 0,746 1,379 0,010 Apsilankymų pas psichiatrus sk./100 gyv. 0,942 0,366 0,325 0,013 Modelio: R2 = 0,281, F = 5,07, p = 0,001 a Svorinė mažiausių kvadratų regresija dėl lokalių populiacijų dydžio skirtumų; Modelis 2: kontroliuotas nedarbo lygio ir apsilankymų pas psichiatrus skaičiaus iškraipantis poveikis; SE, standartinė paklaida.

Moterų savižudybių SMR ir ličio koncentracijos sąsajos Moterų savižudybių SMR statistiškai reikšmingas atvirkštinis ryšys (p = 0,040) su ličio koncentracija geriamajame vandenyje buvo nustatytas tik pasiekus 13,2 µg/l lygį (4.3.6 lentelė).

4.3.6 lentelė. Moterų savižudybių SMR ir ličio koncentracijos sąsajos (dau- gialypės svorinės regresijos analizė) Koeficientaia Nestandartizuoti Standartizuoti p B SE β Modelis 1 (Konstantė) 13,5 0,95 <0,001

Xc [Litis – vidurkis] 0,093 0,079 0,206 0,244 2 Xc [Litis – vidurkis] –0,017 0,007 –0,424 0,019 Modelio: R2 = 0,109, F = 3,1, p = 0,056

96 4.3.6 lentelės tęsinys Koeficientaia Nestandartizuoti Standartizuoti p B SE β Modelis 2 (Konstantė) 18,5 8,1 0,025

Xc [Litis – vidurkis] 0,058 0,075 0,128 0,449 2 Xc [Litis – vidurkis] –0,017 0,008 –0,407 0,040 Nedarbo lygis, proc. 0,256 0,154 0,882 0,103 Apsilankymų pas psichiatrus sk./100 gyv. 0,207 0,076 0,359 0,009 Modelio: R2 = 0,227, F = 4,1, p = 0,004 a Svorinė mažiausių kvadratų regresija dėl populiacijos dydžio skirtumų; Modelis 2: kontroliuotas nedarbo lygio ir apsilankymų pas psichiatrus skaičiaus iškraipantis poveikis; SE, standartinė paklaida.

Apibendrinant, nustatytas ličio koncentracijos geriamajame vandenyje ir SMR nuo savižudybių netiesinis apverstos U formos ryšys. Ličio koncentracijos ir savižudybių SMR sąsaja nebuvo rasta, esant žemesnei negu 15,3 µg/l ličio koncentracijai. Ličio koncentracijos apsauginis poveikis savižudybės rizikai gali būti susijęs su tam tikro lygio ličio koncentracija (>15 μg/l). Tai reiškia, kad ličio koncentracija, įvertinus iškraipantį nedarbo lygio ir apsilankymų pas psichiatrus skaičiaus poveikį, kai kuriose Lietuvos savivaldybėse gali būti veiksnys, įtakojantis mirtingumo rodiklius nuo savižudybių.

4.4. Savižudybių SMR ir sergamumo afektiniais sutrikimais sąsajos su ličio koncentracija Antrame tyrimo etape suformavome išvadą, kad litis gali būti susijęs su mažesniais savižudybių SMR ličio koncentracijai geriamajame vandenyje pasiekus tam tikrą lygį. Ličio koncentracijos mediana buvo lygi 7 μg/l. Atsižvelgiant į tai, analizė buvo pakartota suskirsčius savivaldybes į dvi grupes ličio koncentracijos medianos atžvilgiu, pavadinus jas mažos ir didelės ličio koncentracijos ekspozicijos grupėmis. Abiejų grupių kintamieji palyginti 4.4.1 lentelėje. Tarp šių grupių nenustatyti sociodemografinių rodiklių skirtumai.

97 4.4.1 lentelė. Savivaldybių savižudybių SMR ir sociodemografinių rodiklių pasiskirstymas grupėse ličio koncentracijos medianos atžvilgiu Litis, μg/l Maža (<7,0) Didelė (≥7,0) F (1,51) p N=27 N=26 Savižudybių SMR bendras 42,42±11,27 42,67±11,8 0,006 0,938 Savižudybių SMR vyrų 78,37±24,79 79,75±19,54 0,05 0,824 Savižudybių SMR moterų 13,15±4,45 13,41±5,85 0,03 0,854 Vidutinis gyventojų sk. 42176±64932 36418±21224 0,156b Moterų/vyrų santykisa 1137,07±49,52 1134,62±46,99 0,34 0,854 Nedarbo lygis, proc. 11,18±3,29 11,46±2,56 0,13 0,724 Skyrybų sk./1000 gyv. 3,12±0,52 3,26±0,37 1,28 0,262 Apsilankymų pas 26,86±8,49 28,07±7,94 0,29 0,594 psichiatrus sk./100 gyv. SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); a moterų skaičius tenkantis 1000 vyrų; b Mann-Whitney testas; SN, standartinis nuokrypis.

Sergamumo psichikos ir elgesio sutrikimais rodiklių palyginimas tarp savivaldybių grupių parodė didesnį sergamumą afektiniais sutrikimais (F1,51 = 8,0, p = 0,007) ir bendrą sergamumą psichikos ir elgesio sutrikimais (F1,51 = 20,26; p<0,0001) didelės ličio koncentracijos ekspozicijos grupėje, lyginant su mažos ličio koncentracijos grupe (4.4.2 lentelė). Santykinis sergamumo afektiniais sutrikimais rodiklių skirtumas tarp grupių siekė 26,9 proc., o antidepresantų vartojimo santykinis skirtumas buvo 19,2 proc.

4.4.2 lentelė. Savivaldybių sergamumo rodiklių, bandymų žudytis ir antide- presantų vartojimo sk. pasiskirstymas grupėse ličio koncentracijos medianos atžvilgiu Litis, μg/l Rodikliai, 100 000 gyv. Maža (<7,0) Didelė (≥7,0) F (1,51) p N=27 N=26 Psichikos ir elgesio sutrikimai 2553,90±629,84 3246,75±476,91 20,26 <0,001 (PES) Afektiniai sutrikimai, bendras 393,28±189,41 558,71±234,66 8,0 0,007 Afektiniai sutrikimai, vyrų 205,41±123,64 278,85±141,92 4,11 0,048 Afektiniai sutrikimai, moterų 564,64±253,90 783,12±324,99 7,57 0,001 Nervų sistemos ligos 4364,49±1795,55 4486,37±1289,76 0,08 0,778 Šizoafektiniai sutrikimai 44,98±21,90 54,49±24,33 2,24 0,141

98 4.4.2 lentelės tęsinys Litis, μg/l Rodikliai, 100 000 gyv. Maža (<7,0) Didelė (≥7,0) F (1,51) p N=27 N=26 PES vartojant alkoholį 267,12±109,37 306,43±82,84 2,16 0,148 PES vartojant alkoholį, vyrų 454,99±193,70 500,57±150,90 0,91 0,345 PES vartojant alkoholį, moterų 98,49±42,16 134,18±41,79 9,57 0,003 Bandymų žudytis sk. 57,50±4,75 54,88±4,82 0,006 0,938 Antidepresantų vartojimas 17,89±8,61 22,14±7,42 3,64 0,062 1000 gvy.a PES, psichikos ir elgesio sutrikimai; SN, standartinis nuokrypis; a skaičius DDD/1000 gy- ventojų per dieną (ATC/DDD metodologija, paros dozių skaičius).

4.4.1 pav. Sergamumo afektiniais sutrikimais rodiklių ir ličio koncentracijos ryšys Kairėje – tiesinė, p = 0,004; dešinėje – kubinė, p = 0,020. Vertikali brūkšninė linija – ličio koncentracijos mediana 7 µg/l, horizontali – sergamumo afektiniais sutrikimais rodiklių vidurkis 419/100 000 gyv.

4.4.2 pav. Savižudybių SMR ir ličio koncentracijos ryšys Kairėje – tiesinė (p = 0,212) dešinėje kubinė (p = 0,030). Brūkšninė linija žymi ličio koncentracijos medianą 7 µg/l.

Pateiktose diagramose (4.4.1 pav.), matyti, kad sergamumo afektiniais sutrikimais rodiklių svyravimai siekė vidurkio (419/100 000 gyv.) liniją ties ličio koncentracijos mediana (7 µg/l). Didėjant ličio koncentracijai sergamumo afektiniais sutrikimais rodiklių (4.4.1 pav.) ir savižudybių SMR kitimas (4.4.2 pav.) buvo priešingos krypties. Mažos ličio koncentracijos grupė: daugialypės regresijos analizė Mažos ličio koncentracijos grupėje nebuvo nustatyta ličio koncentracijos sąsaja su savižudybių SMR. Šioje grupėje bendras savižudybių SMR buvo susijęs su nedarbo lygiu (β = 0,494, p = 0,003) ir sergamumo afektiniais sutrikimais rodikliais (β = 0,372, p = 0,022). Šie kintamieji paaiškino daugiau negu pusę (R2 = 52,8 proc., p<0,001) viso bendro savižudybių SMR kitimo. Vyrų savižudybių SMR buvo susijusi su nedarbo lygiu (β = 0,439, p = 0,011) ir sergamumu psichikos ir elgesio sutrikimais (β = 0,397, p = 0,020) (R2 = 0,498, p>0,001). Moterų savižudybių SMR priklausė nuo apsilankymų pas psichiatrus skaičiaus (β = 0,416, p = 0,009) ir nuo sergamumo afektiniais sutrikimais (β = 0,503, p = 0,002) (R2 = 0,509, p<0,001). Didelės ličio koncentracijos grupė: daugialypės regresijos analizė 4.4.3 ir 4.4.4 lentelėse pateikti statistiškai reikšmingi ir parinkti reikš- mingi analizei koreliacijų koeficientai (visa lentelė Annex 2).

100 4.4.3 lentelė. Savižudybių SMR ir sociodemografinių rodiklių koreliacijos didelės ličio koncentracijos grupėje Spearman’o koreliacijos koeficientai Rodikliaia 1 2 3 4 5 1 Litis, µg/l 1 2 Savižudybių SMR, bendras –0,46* 1 3 Savižudybių SMR, vyrų –0,45* 0,93** 1 4 Savižudybių SMR, moterų –0,34 0,67** 0,52** 1 5 Vidutinis gyventojų sk. 0,21 –0,47** –0,39* –0,36 1 6 Moterų/vyrų santykisb 0,42* –0,32 –0,23 –0,30 0,06 7 Nedarbo lygis, proc. –0,24 0,50** 0,41* 0,47* –0,43* a rodiklių numeracija sąlyginė; visa lentelė Annex 2; b moterų skaičius tenkantis 1000 vyrų; SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); *p<0,05; **p<0,01.

Statistiškai reikšmingų koreliacijų tarp bendro savižudybių SMR ir sergamumo PES rodiklių nebuvo nustatyta. Tačiau rasta sąsaja tarp vyrų savižudybių SMR ir moterų sergamumo PES rodiklių, bei su PES vartojant alkoholį rodikliais. Remiantis šiomis koreliacijomis, buvo analizuojami ličio koncentracijos poveikį iškraipantys veiksniai atliekant regresinę analizę.

4.4.4 lentelė. Savižudybių SMR, sociodemografinių ir sergamumo rodiklių koreliacijos didelės ličio koncentracijos grupėje Spearman’o koreliacijos koeficientai Rodikliaia 3 8 10 11 14 15 3 Savižudybių SMR, vyrų 1 8 Skyrybų sk./1000 gyv. 0,09 1 Apsilankymų pas 9 0,14 0,16 psichiatrus sk./100 gyv. 10 Afektiniai sutrikimai –0,19 –0,02 11 Nervų sistemos ligos 0,07 0,41* 0,43* 12 Šizofreniniai sutrikimai 0,19 0,22 0,38 0,44* 13 Psichikos ir elgesio sutrikimai –0,21 0,06 0,63** 0.40* 14 PES vartojant alkoholį, bendras 0,26 0,26 –0,29 –0,18 15 PES vartojant alkoholį, vyrų 0,20 0,28 –0,24 –0,13 0,96** 16 PES vartojant alkoholį, moterų 0,42* 0,31 –0,17 0,07 0,80** 0,71** a rodiklių numeracija sąlyginė; visa lentelė Annex 2; SMR, standartizuotas mirtingumo rodiklis 100 000 gyv. (Europos standartas); PES, psichikos ir elgesio sutrikimai; *p<0,05; **p<0,01.

101 4.4.5–4.4.7 lentelės demonstruoja daugialypės regresijos analizės rezultatus. Pateikti nepriklausomi kintamieji susiję su bendru, vyrų ir moterų sa- vižudybių standartizuoto mirtingumo rodikliais savivaldybėse su didele ličio koncentracija geriamajame vandenyje (virš medianos ≥7 µg/l).

4.4.5 lentelė. Savižudybių SMR ir ličio koncentracijos sąsajos didelės ličio koncentracijos grupėje Koeficientai Nestandartizuoti Standartizuoti t p B SE β Grubus modelis (konstantė) 55,181 5,325 10,363 <0,001 Litis, µg/l –0,643 0,252 –0,462 –2,555 0,017 Modelio: R2 = 0,462, F = 6,529, p = 0,017 Modelis 2 (konstantė) 30,679 11,178 2,745 0,012 Litis, µg/l –0,504 0,236 –0,363 –2,135 0,044 Nedarbo lygis, proc. 1,901 0,781 0,413 2,433 0,023 Modelio: R2 = 0,375, F = 6,894, p = 0,005 Grubus modelis – porinis; 2 modelis – kontroliuojant sociodemografinius ir sergamumo psichikos ir elgesio sutrikimais rodiklius; SE, standartinė paklaida.

4.4.6 lentelė. Moterų savižudybių SMR ir ličio koncentracijos sąsajos didelės ličio koncentracijos grupėje Koeficientai Nestandartizuoti Standartizuoti t p B SE β Grubus modelis (konstantė) 18,008 2,795 6,444 <0,001 Litis, µg/l –0,236 0,132 –0,343 –1,787 0,087 Modelio: R2 = 0,117, F = 3,194, p = 0,087 Modelis 2 (konstantė) 24,215 4,744 5,104 <0,001 Litis, µg/l –0,306 0,116 –0,455 -2,637 0,015 Nervų sistemų ligos –0,003 0,001 –0,560 –2,993 0,007 Šizofreniniai sutrikimai 0,120 0,043 0,500 2,767 0,011 Modelio: R2 = 0,423, F = 5,369, p = 0,006 Grubus modelis – porinis; 2 modelis – kontroliuojant sociodemografinius ir sergamumo psichikos ir elgesio sutrikimais rodiklius; SE, standartinė paklaida.

102 4.4.7 lentelė. Vyrų savižudybių SMR ir ličio koncentracijos sąsajos didelės ličio koncentracijos grupėje Koeficientai Nestandartizuoti Standartizuoti t p B SE β Grubus modelis (konstantė) 99,988 8,863 11,281 <0,001 Litis, µg/l –1,040 0,419 –0,452 –2,484 0,020 Modelio: R2 = 0,204, F = 6,168, p = 0,020 Modelis 2 (konstantė) 73,394 12,758 5,753 <0,001 Litis, µg/l –1,068 0,374 –0,465 –2,855 0,009 PES vartojant alkoholį, moterų 0,202 0,076 0,433 2,659 0,014 Modelio: R2 = 0,391, F = 7,398, p = 0,003 PES, psichikos ir elgesio sutrikimai; Grubus modelis – porinis; 2 modelis – kontroliuojant sociodemografinius ir sergamumo PES rodiklius; SE, standartinė paklaida.

Apibendrinant gautus rezultatus, galime teigti, kad ličio koncentracija geriamajame vandenyje (virš medianos), buvo neigiamai susijusi tiek su bendru, tiek su vyrų ir moterų mirtingumu nuo savižudybės, kontroliuojant pagal sociodemografinius ir sergamumo psichikos bei elgesio sutrikimais rodiklius. Savižudybių SMR yra atvirkščiai susiję su ličio koncentracija geriamajame vandenyje tik savivaldybėse, kuriose yra didesnė ličio koncentracija (viršijanti medianą) ir didesnis afektinių sutrikimų paplitimas.

5. Išvados

1. Lietuvos savivaldybių centrinių vandenviečių geriamajame vandenyje vidutinė ličio koncentracija buvo 11,5 (SN 9,9) µg/l, kitimo intervalas nuo 1,0 iki 39,0 µg/l, mediana – 7,0 (tarpkvartilinis intervalas 3,5–20) µg/l. Ličio koncentracija yra skirtingai pasiskirsčiusi visoje Lietuvoje. Mažiausia ličio koncentracija yra Rytų Lietuvoje, Žemaitijoje ir Šilutėje, didžiausia ličio koncentracija – Vidurio Lietuvos, Šiaurės Lietuvos ir Klaipėdos regionuose. 2. Ličio koncentracija yra netiesiškai susijusi su savižudybių rodikliais. Rastas atvirkštinis ryšys tarp didesnės ličio koncentracijos ir mažesnių mirtingumo nuo savižudybių rodiklių, kontroliuojant sociodemografinių veiksnių iškraipantį poveikį. Ličio koncentracija geriamajame vandenyje reikšmingai neigiamai susijusi su savižudybių mirtingumu tik nuo tam

103 tikros ličio reikšmės (14,5 µg/l). Panašus ryšys rastas tarp ličio koncentracijos ir moterų savižudybių SMR, vyrams toks ryšys nebuvo paste- bėtas. 3. Ličio koncentracija geriamajame vandenyje yra teigiamai susijusi su afektinių sutrikimų paplitimu. Rezultatai rodo, kad afektinių sutrikimų paplitimas yra didesnis savivaldybėse, kuriose ličio koncentracija geriamajame vandenyje yra didesnė negu mediana, palyginti su savivaldy- bėmis, kurių ličio koncentracija yra mažesnė už medianą, ir kurioms būdingos tos pačios sociodemografinės ir psichiatrinės savybės. Savižu- dybių SMR yra atvirkščiai susiję su ličio koncentracija geriamajame vandenyje tik savivaldybėse, kuriose yra didesnė ličio koncentracija (viršijanti medianą) ir didesnis afektinių sutrikimų paplitimas.

6. Rekomendacijos

Didelis savižudybių skaičius išsivysčiusiose šalyse formuoja poreikį rengti naujas psichikos sveikatos strategijas įgyvendinant savižudybių pre- venciją. Remiantis mūsų tyrimo rezultatais ir kitų šalių tyrimais, ličio koncentracija geriamajame vandenyje gali neigiamai paveikti savižudybių rodiklius. Tačiau šiai hipotezei reikia išsamesnių tyrimų, kurie įvertintų mažų ličio dozių veikimo mechanizmus. Šiame tyrime mes neįrodome jokio priežastinio ryšio, tačiau tikimės, kad tyrimo metu rastos sąsajos prisidės prie savižudybių rodiklių mažėjimo ateityje, ir pasitarnaus formuojant psichikos sveikatos politiką. Praktinės rekomendacijos. Tyrimo rezultatai atkreipia dėmesį į mažai analizuojamą ličio poveikį savižudybėms Lietuvoje, nepaisant tvirtų įrody- mų. Šio tyrimo rezultatai pabrėžia būtinybę dalintis informacija su psichikos sveikatos politikos formuotojais ir atstovais apie galimą ličio geriamajame vandenyje antisuicidinį poveikį, akcentuojant jo ryšį su savižudybių rodikliais ir afektinių sutrikimų paplitimu. Sveikatos politikos formuotojams, įgyvendinant savižudybių prevencijos priemones psichikos sveikatos sutrikimais sergantiems asmenims, būtina atkreipti dėmesį į regioninius savižudybių rodiklių netolygumus, sutelkiant dėmesį į savivaldybes kuriose išlieka didžiausias savižudybių skaičius. Mokslinės rekomendacijos. Iki šiol trūksta duomenų, kokia ličio koncentracija geriamajame vandenyje gali suteikti apsauginį poveikį nuo savižu- dybės. Reikalingi tolesni tyrimai, kad suprastume, kaip ličio koncentracija geriamajame vandenyje koreliuoja su ličio koncentracija žmogaus orga- nizme.

104 Remiantis mūsų tyrimo duomenimis, ištirti geriamojo vandens mėginiai gali būti būdas patikrinti šią hipotezę, ypač regionuose, kuriuose pastebimas didesnis afektinių sutrikimų paplitimas ryšyje su savižudybių rodikliais. Papildomi ir išsamūs tyrimai suteiktų naujų žinių vertinant biologinius ličio mechanizmus suicidiniam elgesiui. Gauti tyrimo rezultatai pateikia įžvalgų naujiems tyrimams, ličio ir nervų sistemos ligų ir (arba) kitų su sveikata susijusių rodiklių sąsajų vertinimui. Vertėtų ištirti ličio koncentraciją ne tik centrinėse, bet ir rečiau naudojamose vandenvietėse ar individualiuose šuliniuose, ypač savivaldybėse, kurios pasi- žymi didesniais savižudybių rodikliais. Nepaisant poreikio atlikti daugiau tyrimų, šis tyrimas papildo ličio ir savižudybių sąsajų tyrinėjimų sritį parodydamas jų ryšį su afektiniais sutrikimais, bei poreikį vystyti tolesnius tyrimus individualiame lygyje, kurie padėtų atskleisti priežastinį ryšį ir atsakyti į keliamus klausimus.

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116 PUBLICATIONS ON THE DISSERTATION THEME

1. Liaugaudaite V, Naginiene R, Raskauskiene N, Mickuviene N, Bunevi- cius A, Sher L: Relationship between Lithium Levels in Drinking Water and Suicide Rates: A Nationwide Study in Lithuania. Archives of Suicide Research 2019;0:1–13. DOI: 10.1080/13811118.2019.1674226 2. Liaugaudaite V, Mickuviene N, Raskauskiene N, Naginiene R, Sher L. Lithium levels in the public drinking water supply and risk of suicide: a pilot study. J Trace Elem Med Biol 2017;43:197-201.

PRESENTATIONS ON THE DISSERTATION THEME 1. Liaugaudaitė, V.; Naginienė, R.; Raškauskienė, N.; Mickuvienė, N.; Bunevičius, A.; Sher, L. Inverse relationship between lithium levels in drinking water and suicide rates // European neuropsychopharmacology: Abstracts of the 32nd ECNP Congress: 7-10 September 2019, Copenha- gen (Denmark): Elsevier. ISSN 0924-977X. eISSN 1873-7862. 2019, vol. 29, suppl. 6, p. S401-402, No. P.574. Poster presentation. 2. Liaugaudaitė, Vilma; Raškauskienė, Nijolė; Naginienė, Rima; Micku- vienė, Narseta. Inverted U-shaped relation between natural lithium levels in drinking water and suicide rates //32nd Nordic Congress of Psychiatry: 13-16 June, 2018, Reykjavik (Iceland). Oral presentation. 3. Liaugaudaitė, Vilma; Raškauskienė, Nijolė; Mickuvienė, Narseta. The association between natural lithium levels in drinking water and suicide rates in Lithuania // 1st International doctoral students’ conference “Scien- ce for Health”: book of abstracts: April 13, 2018, Kaunas, Lithuania / Lithuanian university of health sciences. LSMU Department of Research Affairs. Council of LSMU Doctoral Students; [Edited by Indrė Šveikauskaitė]. Kaunas: Lietuvos sveikatos mokslų universiteto Leidybos namai, 2018. ISBN 9789955155300. p. 91-92. Oral presentation. 4. Liaugaudaitė, Vilma; Raškauskienė, Nijolė; Mickuvienė, Narseta. Lithium levels in the public water supply and risk of suicide // European neuropsychopharmacology: Abstracts of the 29th ECNP Congress: 17– 20 September, 2016 Vienna (Austria): Elsevier. ISSN 0924-977X. 2016, vol. 26, suppl. 2, p. S448, No. P.2.e.004. Poster presentation.

117 5. Liaugaudaitė, Vilma; Raškauskienė, Nijolė; Naginienė, Rima; Micku- vienė, Narseta. Savižudybių sąsajos su ličio koncentracija geriamajame vandenyje: pilotinis tyrimas // I-oji nacionalinė mokslinė – praktinė konferencija „Visuomenės sveikata saugiai Lietuvai“: konferencijos tezių knyga: 2016 m. spalio 6 d., Kaunas / Sudarytojai: Ramunė Kalė- dienė, Mindaugas Stankūnas, Vilma Jasiukaitienė, Paulius Vasilavičius, Jurgita Vladičkienė; Lietuvos sveikatos mokslų universiteto Visuomenės sveikatos fakultetas. Kaunas: Lietuvos sveikatos mokslų universitetas, 2016. ISBN 9789955154525. p. 36-36. Oral presentation.

118 119 120 121 122

123 124 125 126 127 128 129 130 131 132 133 134 135

136 ANNEXES

137 Annexe 1

Table 1. The low lithium exposure group: Spearman correlation coefficients between variables 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 1 Lithium 1 2 SMR_T 0.14 1 3 SMR_M 0.12 0.92** 1 4 SMR_W –0.14 0.48* 0.39** 1 5 Population size 0.04 –0.43* –0.41* –0.26 1 6 Women/mena –0.15 –0.39* –0.47* –0.09 0.18 1 7 Unemployment –0.04 0.39* 0.33 0.19 –0.26 0.05 1

1 38 rate 8 Divorce –0.28 –0.17 –0.16 0.14 –0.14 –0.19 0.89 1 9 Visits to –0.04 0.21 0.11 0.19 –0.20 0.22 0.43* –0.13 1 psychiatrist 10 Affective 0.14 0.34 0.34 0.30 –0.23 –0.16 0.12 –0.21 –0.04 1 Disroders 11 Nervous D 0.12 0.06 0.13 –0.11 –0.11 0.13 0.02 –0.31 –0.24 0.28 1 12 SCH 0.13 –0.01 –0.06 0.06 –0.05 0.24 0.29 –0.04 0.19 0.30 0.06 1 13 MBD 0.19 0.32 0.35 0.25 –0.21 –0.03 0.13 –0.38 0.27 0.71** 0.39* 0.27 1 14 MBD/A 0.48* 0.37 0.25 0.06 –0.26 –0.38 0.32 –0.21 0.34 0.29 0.16 0.31 0.49** 1 15 MBD/A-M 0.43* 0.39* 0.26 0.12 –0.26 –0.35 0.36 –0.18 0.35 0.29 0.13 0.32 0.48* 0.99** 1 16 MBD/A-W 0.57** 0.18 0.15 –0.21 –0.21 –0.29 0.09 –0.29 0.27 0.19 0.28 0.27 0.48* 0.78** 0.68** 1 *p<0.05; **p<0.01; D, disease; M, men; W, women; MBD, mental and behavioral disorder; MBD/A, mental and behavioral disorder due to use of alcohol; SCH, schizophrenia; a number of women for 1,000 men; SMR, standardized mortality rate per 100,000 (European standard population). Annexe 2

Table 2. The high lithium exposure group: Spearman correlation coefficients between variables 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 1 Lithium 1 2 SMR_T –0.46* 1 3 SMR_M –0.45* 0.93** 1 4 SMR-W –0.34 0.67** 0.52** 1 5 Population size 0.21 –0.47** –0.39* –0.36 1 6 Women/mena 0.42* –0.32 –0.23 –0.30 0.06 1 7 Unemployment –0.24 0.50** 0.41* 0.47* –0.43* 0.04 1

13 9 rate 8 Divorce rate –0.05 –0.11 0.09 –0.19 0.28 0.26 –0.13 1 9 Visits to 0.13 0.15 0.14 0.22 –0.06 –0.13 0.23 0.16 1 psychiatrist 10 Affective 0.19 –0.15 –0.19 0.002 –0.33 –0.07 0.06 –0.02 0.13 1 Disorders 11 Nervous D –0.28 –0.02 0.07 –0.21 –0.09 –0.12 –0.17 0.41* 0.20 0.43* 1 12 SCH –0.11 0.22 0.19 0.30 –0.22 0.05 0.07 0.22 0.34 0.38 0.44* 1 13 MBD –0.08 –0.19 –0.21 –0.11 –0.34 0.21 0.13 0.06 0.16 0.63** 0.40* 0.23 1 14 MBD/A 0.05 0.16 0.26 0.18 –0.15 0.36 0.26 0.26 0.19 –0.29 –0.18 0.11 0.15 1 15 MBD/A-M 0.09 0.07 0.20 0.14 –0.11 0.33+ 0.17 0.28 0.17 –0.24 –0.13 0.15 0.22 0.96** 1 16 MBD/A-W 0.03 0.36 42* 0.14 –0.20 0.19 0.30 0.31 0.25 –0.17 0.07 0.31 0.10 0.80** 0.71** 1 *p<0.05; **p<0.01; D, disease; T total; M, men; W, women; MBD, mental and behavioral disorder; MBD/A, mental and behavioral disorder due to use of alcohol; SCH, schizophrenia; a number of women for 1,000 men; SMR, standardized mortality rate per 100,000 (European standard population). Annexe 3

Figure 1. Box plot: identified outlier in dataset of the lithium levels in drinking water

140 CURRICULUM VITAE

Name, Surname: Vilma Liaugaudaitė E-mail: [email protected]

Education 2016–2020 Doctoral studies in Public Health, Lithuanian University of Health Science, Kaunas, Lithuania 2009–2012 Master in Management of Public Health, Lithuanian University of Health Science, Kaunas, Lithuania 2005–2009 Bachelor in Public Health, Kaunas University of Medicine, Kaunas, Lithuania

Work experience 2020–2021 Research assistant in Research project “Determinants of quality of life in Lithuanian students: problematic usage of the Internet and neuropsychological profile”. 2017–2020 Research assistant in Research project “Psychosocial suicide risk factors and accessibility of help in the environment of persons’ who committed suicide”. 2015–present Clinic administrator, Hospital of Palangos Klinika of Neuroscience Institute Lithuanian University of Health Sciences, Palanga, Lithuania 2012–present Research assistant, Lithuanian University of Health Science, Lithuania

Teaching experience 2017–2018 Assistant of the Master program “Lifestyle Medicine” for the course “Mental Health and Lifestyle” in Clinical Department of Behavior Medicine Lithuanian University of Health Sciences

Academic awards and travel support 2013 Travel grant from Science Support Fund of Lithuanian University of Health Science 2013 Research scholarship from European College of Neuropsychopharmacology 2014 European College of Neuropsychopharmacology grant for participation in a workshop 2015 European College of Neuropsychopharmacology Seminar Award 2016 European College of Neuropsychopharmacology Research Internship 2016 Travel grant from Science Support Fund of Lithuanian University of Health Science 2016 Research scholarship from European College of Neuropsychopharmacology 2017 Travel grant from Research Council of Lithuania 2018 Travel grant from Research Council of Lithuania 2018 International Doctoral Students' Conference “Science for Health 2018” Oral Presentation Award

141 Research-related training and courses 24-26/04/2015 European College of Neuropsychofarmacology / Baltic Regional Seminar in Neuropsychofarmacology / Kernave (Lithuania) 06-09/03/2014 European College of Neuropsychofarmacology / Workshop / Nice (France) 15-17/05/2013 European College of Neuropsychofarmacology / Baltic Regional Seminar in Neuropsychofarmacology, Baltezers (Latvia)

Scientific visits 19/02/2017-05/03/2017 Internship at the Geha Mental Health Center, Child and Adolescent Department, Petah Tikva (Israel)

Memberships 2019-up to date Junior member of the International Academy of Suicide Research (IASR) 2017-up to date COST Action Member (CA16207 – European Network for Proble- matic Usage of the Internet) (National Representor since 2018) 2014-up to date Member of Lithuanian Society of Biological Psychiatry 2013-up to date Associate member of European College of Neuropsychopharma- cology (ECNP)

142 ACKNOWLEDGEMENTS

Many people deserve thanks and acknowledgment for their efforts and contributions to this research and my development as a researcher. I wish to express my gratitude to all my colleagues for their help during all these years of my work. First of all, I wish to thank Habil. Dr. Robertas Bunevičius, who offered me an opportunity to start my scientific career and in parallel was the first supervisor of my scientific work. His valuable advice and ideas were immeasurable and provided the basis on this thesis. I am grateful to my scientific supervisor Prof. Dr. Nida Žemaitienė, for accepting me as a doctoral student as well as for her support and supervision, especially at the finishing stages of arrangement of the thesis. I am grateful to my scientific consultant Assoc. Prof. Dr. Narseta Mickuvienė, for continuous scientific supervision and consultations. Her helpful and critical comments on my work are very important for further research subjects. I’m extremely grateful to my colleague Nijolė Raškauskienė for her valuable advice on methodological and statistical issues, especially for her professional help in statistical analysis, her time and patience. Her professional knowledge and support were invaluable during my thesis. I would like to say thanks to Habil. Dr. Julija Brožaitienė for her continuous encouragement and support. I would like to acknowledge the co-authors of my scientific papers and my colleagues for remarks, advices, and stimulating discussions. I am grateful to Prof. Habil. Dr. Saulius Šliaupa for sharing his knowledge of geological research, and valuable contribution for the detailed analysis of lithium in drinking water. Throughout the development of the thesis, my work was supported by grants and acknowledgements from the Lithuanian University of Health Sciences, and Research Council of Lithuania Fund. I am grateful to the Laboratory UAB Vandens tyrimai for the analysis of lithium in drinking water and its valuable contribution to this research. Lastly, warmest thanks go to my family and friends, for the support and opportunity to study for many years to reach this important moment of my life.

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