Evaluation of Plasma Levels of Melatonin After Midazolam Or Sodium Thiopental Anesthesia in Children

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J. Pineal Res. 2002; 32:253–256 Copyright Ó Blackwell Munksgaard, 2002 Journal of Pineal Research ISSN 0742-3098 Evaluation of plasma levels of melatonin after midazolam or sodium thiopental anesthesia in children

Mun˜ oz-Hoyos A, Heredia F, Moreno F, Garcı´ a JJ, Molina-Carballo A, Antonio Mun˜ oz-Hoyos1, Escames G, Acun˜ a-Castroviejo D. Evaluation of plasma levels of melatonin Francisco Heredia1, Francisco 2 3 after midazolam or sodium thiopental anesthesia in children. J. Pineal Res. Moreno , Joaquıń Jose´ Garcıá , 1 2002; 32:253–256. Ó Blackwell Munksgaard, 2002 Antonio Molina-Carballo , Germaine Escames4 and Darıó Abstract: Midazolam and sodium thiopental are two commonly used drugs Acun˜ a-Castroviejo4 in anesthesia for minor surgical procedures in children. A relationship exists 1Departamento de Pediatrıá, Facultad de between benzodiazepines (BNZ), barbiturates and melatonin. Whereas these Medicina, Universidad de Granada, Spain; drugs increase pineal melatonin production, the indoleamine amplifies the 2Hospital Comarcal de Baza, Granada, Spain; 3 effects of both BNZ and barbiturates on the central nervous system (CNS). Departamento de Farmacologıá y Fisiologıá, Facultad de Medicina, Universidad de Our purpose was thus to analyze the plasma levels of melatonin before and Zaragoza, Spain; 4Departamento de during midazolam or sodium thiopental anesthesia in children subjected to Fisiologıá, Facultad de Medicina, Universidad ambulatory surgical procedures. Midazolam (0.4 mg/kg) or sodium de Granada, Spain thiopental (5 mg/kg) were administered i.v. to 33 and 32 children (aged between 2 and 14 yr), respectively, and blood samples were taken before and 5, 10 and 20 min after the drugs were administered. Melatonin was measured in plasma by a commercial radioimmunoassay kit previously standardized in our laboratory. The results showed that neither midazolam nor sodium

thiopental anesthesia significantly affected the levels of melatonin studied at Key words: anesthesia, barbiturates, anytime. Significant correlations were found comparing the levels of benzodiazepines, children, oxidative stress melatonin between the different times studied. These results suggest that Address reprint requests to Dr Darıó Acunã- midazolam or sodium thiopental did not affect melatonin production by the Castroviejo, Departamento de Fisiologıá, pineal gland, thus avoiding a possible potentiating effect of the indoleamine Facultad de Medicina, Avda. de Madrid 11, E-18012 Granada, Spain. on the central effects of these drugs during anesthesia. However, the E-mail: [email protected] possibility that changes in melatonin had been masked by the antioxidant Received March 8, 2001; role of the neurohormone are discussed. accepted November 29, 2001.

Introduction These psychopharmacological effects of melatonin suggest an interaction between the indoleamine with brain Midazolam and sodium thiopental are largely used drugs in GABA–BNZ receptor complex [12]. A double-blind, pla- pediatric clinic because of their pharmacological properties. cebo-controlled study comparing melatonin with midazo- Midazolam is better than diazepam for i.v. sedation lam showed that melatonin can be used effectively for because of its faster onset of action, more consistent premedication of adult patients [13]. Nevertheless, no anterograde amnesia, and virtual lack of venous complica- clinical studies delineate the consequences of BNZ and/or tions [1]. Sodium thiopental is a barbiturate also used for barbiturate administration on melatonin levels in children, i.v. anesthesia during brief surgical procedures [2]. although melatonin potentiates the effects of BNZ and Melatonin is an indoleamine produced in several tissues barbiturates in children [4, 8, 14]. Thus, the present study including the pineal gland. Pineal production melatonin is was designed to examine the effects of two common mainly controlled by norepinephrine action on b-adreno- anesthetics used in ambulatory surgery, i.e. midazolam ceptors in the pinealocyte [3]. As the pineal gland is outside and sodium thiopental, on the plasma melatonin levels in the blood–brain barrier, any compound may reach the children under ambulatory surgery. gland with the circulation and may alter its production of melatonin. Melatonin is a benzodiazepine (BNZ)-like drug Methods with hypnotic, anxiolytic, sedative and anticonvulsant properties [4, 5] and its production is suppressed by A total of 65 children, submitted for the University c-aminobutyric acid (GABA) and BNZ administration in Granada hospital for minor surgical procedures, were humans [6–8]. In turn, melatonin administration increased studied. The children have normal psychomotor develop- brain GABA production [9] and modulated the circadian ment, normal clinical and routine biochemical findings, rhythms of brain GABAA and BNZ receptors [10, 11]. absence of obstetrics and/or perinatal antecedents that

253 Munõz-Hoyos et al. might represent neurological risk factors, and absence of familial, neurological or endocrine illness. Information was given and authorization obtained from the parents and from the hospital’s Ethical Committee, and the Code of Ethics of the World Medical Association was observed. The children were randomly divided into two groups: (a) midazolam group, comprising 33 children aged between 2 and 13.5 yr, who received an i.v. injection of 0.4 mg/kg of midazolam, and (b) sodium thiopental group, contained 32 children aged between 2 and 14 yr receiving i.v. injection of 5 mg/kg of sodium thiopental. All children of both groups were classified as class 1 according to the American Society for Anesthesiology classification for physical status. The following anesthetic protocol was performed in all children: (1) atropine, 0.01 mg/kg; (2) sodium thiopental [2,4,6 (1H,3H,5H)-pyrimidinetrione, 5-ethyl-5-(l-methylbu- tyl), Abbot Laboratories, Madrid, Spain], 5 mg/kg (thiopental group) or midazolam (4H-imidazol [1,5-a] [1,4]benzodiazepine, 8-chloro-6-(2-fluorophenyl)-I-Methyl, Roche, Barcelona, Spain), 0.4 mg/kg (midazolam group); (3) vecuronium, 0.1 mg/kg; (4) intubation; (5) alfentanil, starting with 40 lg/kg and successive doses of 15 lg/kg; (6) maintenance with 35% O2N2O; (7) artificial ventilation with VT of 10 mL/kg and a respiratory rate of 16–20; (8) liquid support, between 11 and 20 kg, 4 mL/kg/h; between 21 and 40 kg, 3 mL/kg/h, and above 41 kg, 2.5–3 mL/kg/ h). All drugs were i.v. injected, and the studies were performed between 09:00 and 12:00 hr. Fig. 1. Mean profile of plasma melatonin concentrations before Peripheral blood samples were collected from the ant- (basal) and 5, 10 and 20 min after the administration of midazolan ecubital vein (contralateral to that used for drug injection) (A) or sodium thiopental (B). in groups before (basal) and 5, 10 and 20 min after the drug administration. Blood was centrifuged at 3000 g for 10 min, and plasma aliquots were separated and frozen at for the levels of melatonin between each time of extraction )20°C until assays. Melatonin was determined by a except the correlation for melatonin levels between basal commercial radioimmunoassay (DLD, Hamburg, Ger- and 10 min and between 10 and 20 min in the thiopental many) and a quality control [15] was performed showing group. The coefficient of determination was only significant an intra- and interassay coefficients of variation of 11.3 and in the relation 10–20 min of the midazolam group. 6.3%, respectively. The recovery of added melatonin was 84.4% and the sensitivity of the assay was 0.02 nmol/L. Discussion Data are expressed as mean ± S.E.M. of melatonin values in plasma. Statistics included one-way analysis of Our results show an interesting finding as neither midazo- variance (ANOVA), followed by a Newman–Keuls test, and lam nor sodium thiopental modified the plasma levels of Pearson’s regression and correlation analysis. melatonin. Because of the relationships between BNZ, barbiturates and melatonin, these results were surprising. Results The first consideration is that as we have measured melatonin levels up to 20 min after drug administration, The plasma levels of melatonin before and after midazolam we cannot know if the levels of the neurohormone may administration did not show significant changes (Fig. 1A). change after this time. Circulating melatonin decreases Before the administration of this BNZ, the melatonin during development, but both prepubertal and pubertal concentration was 0.064 ± 0.005 nmol/L. At 5, 10 and children have similar patterns of melatonin production [16, 20 min after anesthesia, the values of melatonin were 17], with a half-life about 0.67–0.78 hr [18]. These data, 0.073 ± 0.007, 0.063 ± 0.005, and 0.068 ± 0.005 nmol/L, with the fact that melatonin produced by the pineal is not respectively. The concentrations of plasma melatonin stored in the gland but it is quickly released to the corresponding to the thiopental group were also without circulation after its synthesis, made it unlikely that changes significant changes during anesthesia (Fig. 1B). In fact, in melatonin levels were not detected during the time of the melatonin levels were 0.0719 ± 0.006 before anesthesia, study. and 0.068 ± 0.006, 0.079 ± 0.007, and 0.073 ± 0.00 The relationships between melatonin, GABA and BNZ nmol/L at 5, 10 and 20 min, respectively, after thiopental are largely known. Melatonin regulates both brain GABA administration. and BNZ binding sites and increases brain GABA levels, The results of the correlation analyses are shown in which may explain its BNZ-like actions [5, 10, 11]. In turn, Table 1. In both groups there was a significant correlation BNZ inhibit pineal production of melatonin and decrease

254 Melatonin and children anaesthesia

Table 1. Correlation analysis between melatonin levels at each measured time Midazolam group Sodium thiopental group Times (min) rR r R

Basal/5 0.56*** 0.31 NS 0.44** 0.19 NS Basal/10 0.48** 0.23 NS 0.31 NS 0.09 NS Basal/20 0.48** 0.23 NS 0.40* 0.16 NS 5/10 0.53** 0.28 NS 0.51** 0.26 NS 5/20 0.54** 0.29 NS 0.53** 0.28 NS 10/20 0.59*** 0.35* 0.34 NS 0.11 NS

*P < 0.05; **P < 0.01; ***P < 0.001; NS ¼ not significant; r ¼ Pearson’s coefficient of correlation; R (r squared) ¼ coefficient of determination. brain melatonin binding sites, an effect that may be oxidative attack [30, 31]. A consequence of the antioxidant counteracted by administration of the indoleamine [19]. ability of melatonin is that during anesthesia increasing These interactions between melatonin and BNZ have been levels of the neurohormone could be not detected because used for reversal BNZ tolerance. Human pineal gland has they are masked by the rapid oxidative metabolism of the both central and mitochondrial BNZ receptors [20], and it indoleamine. Thus, the possibility that during midazolam was reported that BNZ decrease melatonin production [7] or sodium thiopental anesthesia significant changes in inhibiting N-acetyl-5-methoxytryptamine activity (NAT), melatonin production as a protective mechanism against the key enzyme for its synthesis. oxidative damage should be further examined. A comparative study with melatonin and midazolam in adults showed a preoperative anxiolysis and sedation by Acknowledgements the former compatible with its use for premedication [21]. It seems that at the dose used in our study, midazolam This work was partially supported by the Junta de does not induce an increase in melatonin synthesis by the Andalucı´ a (Grupos de Investigacio´ n CTS-101 and CTS- pineal gland, which has two important consequences for 190) and by the Hospital Universitario San Cecilio of the children: (a) there is no more melatonin that could Granada. potentiate the effect of the BNZ on the CNS, delaying the awake time for the children, and (b) there are no changes References in the circadian rhythm of the neurohormone that could alter the synchronization of the melatonin-dependent 1. DUNDEE JW, HALLIDAY NJ, HARPER KW et al. Midazolan: a rhythms. Barbiturates act on the GABAA–BNZ receptor review of its pharmacological properties and therapeutic use. complex increasing GABAergic neurotransmission [22]. Drugs 1984; 28:519–543. Some studies suggest that barbiturates increase pineal 2. FERRENDELLI JA. Pharmacology of antiepileptic drugs. Epi- NAT levels and activity, thus stimulating melatonin lepsia 1987; 28:14–16. synthesis [23]. In turn, the indoleamine potentiates the 3. REITER RJ. Pineal melatonin: cell biology of its synthesis and effects of barbiturates in brain [4] and thus, a feedback of its physiological interactions. Endocr Rev 1991; 12:151–180. regulation between melatonin and barbiturates can be 4. MOLINA-CARBALLO A, MUN˜ OZ-HOYOS A, REITER RJ et al. proposed. The lack of changes in melatonin following Utility of high doses of melatonin as adjunctive anticonvulsant thiopental anesthesia may account also for the dose of the therapy in a child with severe myoclonic experience: two years’ experience. J Pineal Res 1997; 23:97–105. drug used, and yield the same considerations as for 5. NARANJO-RODRI´ GUEZ EB, ORTIZ OSORNIO A, HERNA´ NDEZ- midazolam anesthesia. AVITIA V et al. Anxiolytic-like actions of melatonin, 5-meth- A last consideration should be done in the light of the oxytryptophol, 5-hydroxytryptophol and benzodiazepines on a surgical-induced oxidative stress. Surgical trauma intensi- conflict procedure. Prog Neuro-Psychopharmacol Biol Psy- fies the oxidative stress by generating reactive oxygen chiat 2000; 24:117–129. species (ROS) and diminishing the endogenous defense 6. ROSENSTEIN RE, CHULUYAN HE, PEREYRA EN et al. Release against ROS attack. Thus, antioxidant protection during and effect of gamma-aminobutyric acid (GABA) on rat pineal the perioperative period may be of great importance. melatonin production in vitro. Cell Mol Neurobiol 1989; Midazolam and sodium thiopental are not good antioxi- 9:207–219. dants. Midazolam is a weak antioxidant because it only 7. MCINTYRE I, NORMAN TR, BURROWS GD et al. Alterations to suppresses superoxide anion and reduces mitochondrial plasma melatonin and cortisol after evening alprazolam ROS at concentrations far beyond clinical relevance [24, administration in humans. Chronobiol Int 1993; 10:205–213. 25]. Sodium thiopental protects the neurons from oxidative 8. MONTELEONE P,FORZIATI D, ORAZZO C et al. Preliminary stress, but it spontaneously generates hydroxyl radical observations on the suppression of nocturnal plasma melatonin inducing lipid peroxidation [26]. Melatonin scavenges ROS levels by short-term administration of diazepam in humans. including superoxide anion, hydroxyl radical and hydrogen J Pineal Res 1989; 6:253–258. peroxide [27–29]. Melatonin also increases glutathione 9. ROSENSTEIN RE, CARDINALI DP. Melatonin increases in vivo levels and the activity of enzymes involved in redox GABA accumulation in rat hypothalamus, cerebellum, cereb- homeostasis, protecting mitochondria and cell against ral cortex and pineal gland. Brain Res 1986; 398:403–406.

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