Beyond the Cholinergic Crisis Galle Medical Association Oration 2015

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Beyond the cholinergic crisis Galle Medical Association Oration 2015

Jayasinghe S S Department of Pharmacology, Faculty of Medicine, University of Ruhuna, Galle, Sri Lanka. South Asian Clinical Toxicology Research Collaboration, Department of Medicine, Faculty of Medicine, University of Peradeniya, Peradeniya, Sri Lanka.

Correspondence: Dr. Sudheera S Jayasinghe e-mail: [email protected]

ABSTRACT Introduction: Organophosphates (OP) are the most frequently involved pesticides in acute poisoning. In Sri Lanka it has been ranked as the sixth or seventh leading cause of hospital deaths for many years. Neurotoxic effects of acute OP have been hitherto under-explored. The aims of the studies were to assess the effects of acute OP poisoning on somatic, autonomic nerves, neuromuscular junction (NMJ), brain stem and cognitive function. Methods: Patients following self-ingestion of OP were recruited to cohort studies to evaluated the function of somatic, autonomic nerves, NMJ, brain stem and cognition. Motor and sensory nerve function was tested with nerve conduction studies. Cardiovascular reflexes based autonomic function tests and sympathetic skin response (SSR) was used to evaluate autonomic function. NMJ function was assessed with slow repetitive supramaximal stimulation of the median nerve of the dominant upper limb. Brain stem function and cognitive function were assessed with Brain Stem Evoked Response Audiometry (BERA) and Mini Mental State Examination (MMSE) respectively. The data of the patients were compared with age, gender and occupation matched controls. Results: There were 60-70 patients and equal number of controls in each study. Motor nerve conduction velocity, amplitude and area of compound muscle action potential on distal stimulation, sensory nerve conduction velocity and F-wave occurrence were significantly reduced. At one week the significant impairment in autonomic function were change of diastolic blood pressure 3 min after standing, heart rate variation during deep breathing (HR-DB), SSR-amplitude and post-void urine volume. All except HR-DB were reversed at six weeks. No significant impairment of NMJ function, BERA and MMSE were noted. Conclusions: Sub clinical somatic and autonomic nerve dysfunction was observed. There were no strong evidence of long term effects on NMJ, brain stem and higher function.

Introduction annually (2). In Sri Lanka, the majority of poisoning Organophosphate (OP) compounds have become the cases are self inflicted and 77% of the cases are in the most widely used pesticides for agricultural-pests age range of 11-30 years (3). OP and carbamate throughout the world from the 1980s and the risk of compounds were involved in 74% of pesticide acute and sub acute toxicity is high in humans (1). poisoning (4). Acute pesticide poisoning is a major health problem OP bind to the esteratic site on the acetylcholine especially in developing countries. It was estimated esterase (AChE) molecule, phosphorylates the that one million serious, unintentional poisonings enzyme, and lead to inhibition of its action (5). The occurred and an additional two million people were binding between the esteratic site on the enzyme and hospitalized for attempted suicide with pesticides the phosphorus atom is stable and takes hours or

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weeks to break off, depending on the compound the effects of OP on nervous system including central involved. Studies have shown that a phenomenon of and peripheral nervous systems. Further, it will enzyme aging occurs which involves cleavage of a describe the function of peripheral nervous system radical from the inhibited enzyme, making it including somatic (motor and sensory), autonomic resistant to reactivation. The net result is the (sympathetic and parasympathetic), the junction accumulation of excess acetylcholine (ACh) at the between nerve and muscle, brain and higher cholinergic nerve endings all over the body resulting functions of the brain. in the characteristic clinical manifestations. Following inhibition, recovery of this enzyme occurs at a rate of about one percent per day (5). Restoration Methods of AChE levels occurs by spontaneous or induced Cohort studies were conducted with matched reactivation of the enzyme and by new enzyme controls with the approval of the Ethical Review synthesis (5). Committee, Faculty of Medicine, University of Accumulated acetylcholine at nerve endings initially Ruhuna, Sri Lanka. Informed written consent was stimulates and eventually leads to exhaustion of obtained from the patients and the controls. All cholinergic synapses, resulting in neuromuscular clinical investigations were conducted according to junction (NMJ) dysfunction (5). the principles expressed in the Declaration of Helsinki. The problems occurring after OP poisoning have been identified to some extent. There are three well The patients with self-ingestion of OP were recruited defined syndromes. These are acute cholinergic from a tertiary care hospital and a secondary care crisis, intermediate syndrome, organophosphate hospital in the Southern Province of Sri Lanka induced delayed polyneuropathy (OPIDN) and between June 2008 and September 2009. At the time chronic organophosphate induced neuropsychiatric of recruitment to the study, subjects either had disorders (COPIND) (5). features of the cholinergic syndrome or had been given atropine to counteract cholinergic syndrome in Acute cholinergic crisis occurs due to accumulation the peripheral units and then transferred to the of Ach, hence over activity of the nervous system. As collaborating hospitals. a result the person with poisoning develops muscle twitching, narrowing of air ways, increased OP poisoning was confirmed by the history from the bronchial secretion, slowing of heart rate and low patient and/or accompanying person, the cholinergic blood pressure. This cholinergic crisis is life features and plasma cholinesterase activity (ChE). threatening. Therefore the patient needs to be The controls were recruited from the persons managed immediately to secure the life. Even though accompanying the patients to the tertiary care the patient is recovered from cholinergic crisis, the hospital. Age, gender and occupation matched problems are not over. healthy volunteers who did not have a history of Some patients develop muscle weakness, especially acute pesticide exposure were recruited within one the proximal muscles within four days of ingestion. month of the recruitment of the respective case. Age This entity is called the intermediate syndrome. The of the controls was matched to ±3 years of the intermediate syndrome was first described and patients. published by two Sri Lankans Prof. Nimal Subjects with features of peripheral neuropathy, Senanayaka and Dr. Lakshman Karaliadda in 1987 diabetes mellitus or those who were on long term and they coined the term intermediate syndrome. medications, were excluded. However problems are not yet over. There are things Motor nerve conduction studies (MNCS), sensory happening beyond the cholinergic crisis. OP can also nerve conduction studies (SNCS), F-wave studies, cause polyneuropathy rarely. This occurs one to three autonomic function tests, slow repetitive nerve weeks following poisoning. Patients complain of stimulation (RNS), brain stem auditory evoked parasthesis and calf pain. Deep tendon jerks may be response (BAER) and Mini Mental State absent. Examination (MMSE) were performed at the time Therefore it is worth to look into what happens of discharge from the ward (the first assessment) beyond the cholinergic crisis. This paper overview and at six weeks (the second assessment) following

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acute exposure to OP. All neurophysiological For the ulnar MNCS the recording electrode was investigations done on the patients were carried out placed on the mid portion of the abductor digiti on the controls. The room temperature of the quinti, the reference electrode was placed over the neurophysiology laboratory was maintained at 250C. proximal phalanx of the fifth digit and the ground The patients were assessed twice to explore the acute electrode was attached between recording electrode and subsequent effects on peripheral nerves. The and the stimulating probe. The nerve was stimulated earliest possible time to assess the patients was at the on the palmar aspect of wrist and just distal to the time of discharge and the earliest possible time for osseous groove in the posterior aspect of the medial the second assessment with the least drop outs was at epicondyle of the humerus, i.e., the ulnar groove. the sixth week. For the common peroneal MNCS the recording The Neuropack MEB-9400A/K EMG/EP Measuring electrode was placed over the extensor digitorum S y s t e m ( N i h o n K o d e n ) w a s u s e d f o r brevis muscle, the reference electrode was placed electrophysiological assessment. over the fifth toe and the ground electrode between the recording electrode and the stimulating probe. The nerve was stimulated on the lateral aspect of the SNCS popliteal fossa just medial to the insertion of the tendon of the biceps femoris and along the SNCS were performed on median and ulnar nerves anterolateral surface of the fibula, 3 to 4 cm distal on both sides. The orthodromic method was used. to the proximal tip of the fibular head. Stimulating ring electrodes were placed on the second digit for median SNCS and on the fourth digit Motor nerve conduction velocity (MNCV), for ulnar SNCS. The ring cathode was placed around amplitude and area of the compound muscle action the digit near the metacarpophalangeal joint and the potential (CMAP) on distal stimulation were ring anode around the digit near the distal recorded. interphalangeal joint. A recording electrode was placed over the respective nerve on the anterior aspect of the wrist. Ground electrode was placed F-wave studies between the stimulating and the recording F-wave studies were performed on median, ulnar and electrodes. The nerve was stimulated with tibial nerves. Electrodes were placed as for MNCS supramaximal electrical stimulus. on median and ulnar nerves. For the tibial F-wave Sensory nerve conduction velocities (SNCV) and the studies, the recording, the reference and the ground amplitude of the complex were recorded. electrodes were placed over the abductor hallucis muscle between the great toe and between the recording electrode and the stimulating probe MNCS respectively. The nerve was stimulated at a point slightly posterior and proximal to the medial MNCS were performed on median, ulnar and malleolus. Sixteen stimulations were analyzed, common peroneal nerves on both sides. Surface percentage of F-wave occurrence and minimum electrodes were used. The nerve was stimulated with reproducible F-wave latency were recorded. supramaximal stimulation. For the median MNCS, the recording electrode was placed over the abductor pollicis brevis, the Autonomic function tests reference electrode was placed over the proximal R-R interval based autonomic function tests (heart phalanx of the thumb and the ground electrode was rate response to standing, heart rate variation during attached between recording electrode and the deep breathing and heart rate response to Valsalva stimulating probe. The stimulation was given on the manoeuvre), blood pressure response to standing, palmar aspect of midwrist and at the elbow blood pressure response to sustained handgrip, (antecubital fossa) just medial to the palpable sympathetic skin response and residual urine volume brachial artery. The cathode was placed distal to the were assessed. anode.

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Heart rate response to standing Due to muscle contraction, sympathetic activity and Participants were asked to stand up unaided from vasoconstriction should lead to a rise in blood lying down position as quickly as possible when pressure. The diastolic blood pressure at the end of performing heart rate response to standing. The heart the effort should be at least 16 mmHg higher than rate response was expressed by the 30:15 ratio; ratio before the manoeuver. An increase in the diastolic of longest R-R interval around the 30th beat after blood pressure by 10 mmHg or less is considered as standing to shortest R-R interval around the 15th beat abnormal (8,9). after standing (6,8). Sympathetic skin response (SSR) Heart rate response to deep breathing Sympathetic skin response measures the skin To perform heart rate variation during deep potential which is evoked by stimulation (electric, breathing, participants were asked to breathe deeply auditory and visual). This potential shows the and evenly at 6 breaths per minutes (5 seconds in; 5 functioning of the sympathetic nerves of the sweat second out) for three cycles (30 seconds). Maximum glands (10). Participant's skin was cleaned where the and minimum R-R intervals were analyzed during electrodes were attached to remove any moisture and each cycle and converted to beats per minute. gel from the skin. The active electrode was pasted on Greatest heart rate differences (expiratory- the center of the palm of the dominant hand. The inspiratory difference; E-I difference) during each reference electrode was pasted on the center of the cycle were calculated and the three differences back of the same hand. The ground electrode was averaged (8,9). attached between the active and the stimulating electrodes. The stimulating electrode was placed on the median nerve of the recording wrist and Heart rate response to Valsalva manoeuvre stimulated with the intensity of 25 mA. Six stimulations were administered with at least 30 sec During the Valsalva manoeuvre, the participant intervals between each stimulation. The latency was breathed in to a mouth piece connected to a modified measured from the onset of stimulus artifact to the sphygmomanometer and maintained an expiratory onset of negative deflection. The amplitude was pressure of 40 mmHg for 15 seconds. The ratio of the measured from the baseline to negative or positive longest R-R interval within 20 beats of ending the peak whichever was the highest. Average latency test to the shortest interval during the test was and maximum amplitude were recorded. analyzed. The test was performed three times and the ratios from three Valsalva attempts averaged (8,9). Residual urine volume Blood pressure response to standing Post void residual urine volume was measured ultrasonically after a voluntary void (11,12). Blood pressure was measured while the participant was lying down quietly and around three minutes after standing (9) by using an OMRON SEM-1 Exercise modified slow repetitive supramaximal automatic blood pressure monitor that uses the stimulation oscillometric method of blood pressure measurement. The monitor detects blood movement Exercise modified slow repetitive supramaximal through the brachial artery and converts the stimulation (RNS) was used to assess NMJ function movements into a digital reading. on the median nerve of the dominant upper limb. RNS was carried out administering 10 stimulations at a rate of 3Hz with a filter setting of 20 Hz – 5 kHz. Blood pressure response to sustained handgrip Forearm and hand were fixed by straps to restrict In blood pressure response to sustained hand grip, the movements and to ensure that contraction of the participant was asked to press a hand dynamometer abductor pollicis brevis was isometric. Recording with full strength. Then the hand grip was maintained electrodes were placed over the abducter pollicis at 30% of the maximum strength for five minutes. brevis, the reference electrode was pasted over the

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proximal phalanx of the thumb and the ground - percentage change in amplitude between the electrode was attached between the recording fourth and the first action potential of the third electrode and the stimulating probe (13). Laboratory train of stimuli ((C1-C4)/C1 × 100) 0 temperature was maintained at 25 C during the test. - percentage change in amplitude between the The nerve was stimulated at the wrist at rest- the first fifth and the first action potential of the third train train (A), immediately after 30 seconds maximal of stimuli ((C1-C5)/C1 × 100) isometric exercise - the second train (B), and two A decline of compound muscle action potential minutes after the exercise - the third train (C) (14). (CMAP) amplitude up to 8% with RNS is considered Participants were asked to abduct the thumb against as normal (15). fixed resistance for the isometric exercise (14). The post exercise facilitation and the post exercise The post exercise facilitation was assessed by exhaustion were calculated after the isometric calculating the ratio of the amplitude of the first exercise (13,14). action potential in the second train to the amplitude of the first action potential of the first train (B1/A1) The decrement responses were quantified by (13,14) and the post exercise exhaustion by calculating the percentage change in amplitude as calculating the ratio of the amplitude of the first follows: (15) action potential of the third train to the amplitude of

·At rest - the first train of stimuli (A) the first action potential of the first train (C1/A1) (14). - percentage change in amplitude between the second and the first action potential of the first Brain Stem Auditory Evoked Response (BAER) train of stimuli ((A1-A2) /A1 × 100) BAER is an electrophysiological method that can be - percentage change in amplitude between the used for neurotoxicity testing of auditory function fourth and the first action potential of the first (16). A computer based Neuropack S EMG/EP train of stimuli ((A -A )/A × 100) 1 1 4 1 measuring system MEB-9400 (Nihon Kohden) was - percentage change in amplitude between the fifth used for the BAER. A piece of cotton moistened with and the first action potential of the first train of alcohol was used to clean the patient's skin where the

stimuli ((A1-A5)/A1 × 100) electrodes were attached and the skin rubbed with dry gauze to remove any moisture and gel from the skin. Recording electrodes (active and reference) · Immediately after 30 seconds maximal isometric and grounding electrodes were attached on the exercise - the second train of stimuli (B) patient's head with Elefix paste. The patient was kept - percentage change in amplitude between the on a chair relaxed with eyes closed. To avoid EMG of second and the first action potential of the second the neck, patients were asked not to move their head.

train of stimuli ((B1-B2)/B1 × 100) To avoid EMG of the chin, patient was asked to open his / her mouth slightly. The headphones were kept - percentage change in amplitude between the on the head. The hi-cut and the low-cut filters were fourth and the first action potential of the second set to 3kHz and 100Hz respectively. The skin- train of stimuli ((B -B )/B × 100) 1 4 1 electrode contact impedance was kept below 5kÙ. - percentage change in amplitude between the fifth The stimulation intensity was 90dB and 1000 and the first action potential of the second train of stimulations were averaged. Latency was measured stimuli ((B1-B5)/B1 × 100) in msec. Interpeak latencies of I-III, III-V and I-V were measured (16,17). ·Two minutes after the exercise - the third train of stimuli (C) Mini Mental State Examination - percentage change in amplitude between the Cognitive function was assessed only if the second and the first action potential of the third educational level of the participant was at least up to train of stimuli ((C1-C2)/C1 × 100) year five. People who were suffering from dementia, psychiatric illness and those on medication which

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have the potential to alter cognitive function Statistical analysis (anticonvulsants; vigabatrin, gabapentin and Graph Pad Prism 4 and Social Package of Statistical lamotrigine, minor tranquilizers; diazepam, and Software were used for the statistical analysis. antihypertensive like methyldopa) were excluded Normal distribution of the data was tested with from the study. Cognitive function was assessed Kolmogorov-Smirnov test. Data which had no once the patient had recovered from the acute normal distribution were analyzed with non- cholinergic crisis. parametric tests. The paired T-test was used to A validated Sinhalese translation of the MMSE was compare the results of the first and the second used with a few modifications (18). Table 1 assessment and the unpaired T-test was used to illustrates the modifications made in the MMSE compare the results of the patients and the controls. administered in our study which reflected our need to Correlation of neurophysiological indices with use this in the community. A total score of less than potential confounders was analyzed with 24 out of 30 was considered as cognitive impairment. Spearman’s correlations. Multiple liner regression No response or refusal to perform any item was model was used to adjust for potential confounders. scored as zero.

Results Estimation of plasma ChE activity From a total of 163 acute OP poisoning admissions Generally the term acetylcholinesterase activity is to the collaborating hospitals, 60 -70 patients referred to red blood cell acetylcholinesterase or participated for each study. Mean (SD) GCS on acetylcholinesterase at the nerve tissue. However we admission was 14 (2). All patients received atropine, analyzed ChE (ChE = acetylcholinesterase (AChE) 54 patients received pralidoxime. plus butyrylcholinesterase (BChE)) activity in plasma. The modified Ellman method developed by Plasma ChE activity at four and/or twelve hours Worek F et al. (1999) was used to estimate ChE after the exposure was available in 33 patients. The activity in plasma (19). Plasma samples were median (inter quartile range) of plasma ChE activity obtained from EDTA blood after centrifugation (10 at four and twelve hours was 790 (146 - 2598) min, 500 * g) and stored in 1ml aliquots at -800C until µmol/l/min and 431 (136 - 3068) µmol/l/min analysis. respectively. Prior to analysis the thawed samples were kept on ice None of the patients or the controls had diabetes until analysis. Preparation of inhibited mellitus. The mean HbA1C of patients and the cholinesterase were made by incubating plasma controls were 5.4 ± 0.5% and 5.7 ± 0.6%. samples with PX-ethyl, PX-methyl and obidoxime for 15 min at 370C followed by immediate dilution of the samples (1:100 in diluting reagent) and Function of peripheral nerves freezing. The neurophysiological assessments and the results The activity of cholinesterase was measured with of SNCS, MNCS and F-wave studies are shown in a thermostatted filter photometer (Model 1101 M, the Table 1. Eppendorf, Hamburg, Germany) at 436 nm and 370C using polystyrol cuvets. Impairment of peripheral nerve function was Cholinesterase enzyme activity was calculated by observed at both occasions in the cases compared to using the equation, the controls. In the first assessment these were significant for MNCV of median, ulnar and common Sample(mE / min) -Blank(mE / min) Activity(mmol / l / min) = peroneal nerves, SNCV and CMAP amplitude of 10.6 ulnar nerve and F wave occurrence of median, ulnar and tibial nerves. However no abnormality was The analysis was done at Walther Straub Institute for detected when compared to the standard cut-off Pharmacology and Toxicology, Munich, Germany. values for normal MNCS and SNCS except F-wave occurrence.

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In the second assessment significant worsening of When the number of patients with abnormal peripheral nerve function was seen in common autonomic function test was compared with the peroneal CMAP-area and reduction of tibial F-wave number of controls with abnormal autonomic latency compared to the first assessment. MNCV of function tests (by Chi-Square test) in relation to median and common peroneal nerves, amplitude of standard cut-off values of autonomic function tests, ulnar nerve CMAP on distal stimulation and F-wave no statistically significant difference was observed. occurrence of tibial nerve reversed to normal at the Sixteen patients were not able to perform blood second assessment. pressure response to sustained hand grip due to Seventeen patients were admitted to Intensive Care painful cannula site on their dominant hand. Twenty- Unit. Among them seven patients died. Four of the four patients were not able to complete the test. 10 (40%) survivors underwent neurophysiological Twenty-one patients in the second assessment and assessment at median (range) of 24 (6-40) days after 15 controls were also not able to complete the test. the exposure. Participants (26 in the first assessment, 30 in the second assessment and 55 in the controls) who could Correlation of neurophysiological indices with complete the test were within the normal limits. potential confounders was looked into. None of the However odds ratio of the number of patients who neurophysiological indices significantly correlate were able to complete the test in the first assessment with plasma ChE activity. However MNCV of ulnar; (vs controls) and the second assessment (vs controls) SNCV of median and ulnar, MNCV of ulnar; MNCV were 3.4 and 2.3 respectively. of median, ulnar and tibial F-wave occurrence showed significant negative Spearman’s correlations The complex of SSR was almost flat in seven with smoking habits, PAM therapy and alcohol patients in the first assessment and in one patient in consumption respectively. the second assessment. None of the controls showed When multiple liner regression models were used flat SSR complex. to adjust for potential confounders, effects on SNCV The urinary bladder of most patients was of median nerve, area of CMAP of median nerve catheterized. The catheter was removed only at the and F-wave latency of tibial nerve with prlidoxime time of discharge from the hospital. Hence residual therapy and the area of CMAP of ulnar nerve with the urine volume was measured in 27 patients on type of OP ingested showed statistical significance. discharge and 24 matched controls (Table 2). To determine whether any multicollinearity were In the correlation matrix, it was observed that none of present, and to understand whether there is a strong the bivariate correlations are highly correlated linear association between each predictor variable except alcohol consumption and smoking habits, and all other remaining predictors, the Variance GCS on admission and smoking habits. To determine Inflation Factor (VIF) and condition indices were whether any multicollinearity were present, and examined. None of the VIF exceeds 10 and a to understand whether there is a strong linear condition index exceeds 30 (20). association between each predictor variable and all other remaining predictors, the Variance Inflation Factor (VIF) was examined. None of the VIF Function of autonomic nerves exceeds 10. Statistically significant autonomic dysfunction was Adjusted multiple linear regression revealed seen in the first assessment of the patients compared significant effects of alcohol consumption on to the controls in change of diastolic blood pressure change of SBP 3 min after standing (B = 4.4, SE = (DBP) 3 minutes after standing, HR-DB, SSR- 1.6, p = 0.009) and size of the pupil (B = -0.6, SE = amplitude, SSR-latency and post-void urine volume 0.3, p = 0.04); and the effects of smoking on residual (Table 2). At six weeks recovery of autonomic urine volume (B = 11.8, SE = 5.1, p = 0.03) and dysfunction was observed except in HR-DB size of the pupil (B = 0.5, SE = 0.2, p = 0.04) (21). (Table 2).

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m s s l e

s o

g - )

e r n t s s

s l t

a s n

7 7 o n 3 h a o

‡ 3 9 1 . 0 0 r . . e c - . . t 1

5 5 i 0 C 0 0 t 0 - 0 - n - ) n - - a o a d p c e e ( l i M a t

2 -

(

8 -

- )

3 7

l I . s s o o 8 5 o 6

l e 0 t t t o 6 - t

C t 2 1

3 2 6 2

o v n

o 6 2 r t o o e 1 2 4 7 0 e t t s 1 o o . . . . t t l i

0 0 % 1 t 3 t

t . . n 0

0 0 0 0 .

5 5 . 4 6 a n 5 0 0 . s o 9 1 . . 8 9 - - 1 e p 0 c 5 v 2 1 2 - . (

- - m 0 s

l s t s o e a r e

g t

- t s )

n n s s n s

l a t

o a a

4 0 ‡

‡ o h n ‡ ‡ 6 c ‡ C 0 1 7 . ‡ r i c 6 e . . 6 . 7 t

5 . f i 2 1 - i t 0 0 9 0 1 n - n 4 - - - a n o a p g c e i ( s

M ‡

† p ,

) ) † † ) ) ) ) u ) )

e 4

) 8 2 8 2 o - 7 0 ) g r

3 2 3 9 5 0 1 . 0 . . . n g o o

1 7

t t 0 1 0 0 0 a (

( (

( ( ( r t

o = 2 3

r e - 1 7 4 9 5 1 t 0 l . 4 ( n - 2 3 6 2 (

i n

( 3 . . 5 . t

r o 1 1 1 1 5 - 7 a n - C u o i q r t

† t ) ) ) ) e c - )

n ) 9 6 9 6 n

7 n o e d

6 1 3 6 ) 9 i 1 t . . . . u

n ( o 8 1 5 f m

†

t 0 0 0 0 ( (

o )

s - (

( ( (

n 1 c

c 1 = s 4 2 6 a -

i e - 1 7 4 9 e i ( 2 . 9 ( n

s S 2 6 9 d

m 3 . . 5 . s 6 s e 3 - t a 1 1 1 0 o - n m n

e †

t † t † † t ) ) ) ) ) ) a p )

u n 0 0

) 1 4 9 4 e P

1 ) a e c

1 1

1 3 3 4 3 6

. 2 . . . t x o f 2 6 m s o o t 1 1 0 0 0 (

e r

t t o s ( ( ( ( (

= s ) 8 9 t

3 0 F - 1 2 1 9 1 e - . 3 ( n ( D s n 2 ( 3 5 5

4 . . 6 . s e S 2 3 a 4 1 1 1 0 ( - 2

- m n s a

s e g e s n m i

d ) n

i A

n n

r i

a e e e t : t r t

m s f

a f / n 2

)

g s e a

o l o s t

e i s n t n e t

l i a n m i n i r u c e e (

b l o

r s u † n m ) m a a

p b e n d u 3 )

g s ( 3 )

T V f o

m e n P H p V g P c

o R H s i l

o n B

i ) o m B e m i i t o i n t s m ( r m t S D h

i a

v ) m a

t i o a k ( e e r f f m m e r r a

n m ( r (

S † d o o

e n a

o u a g g i r u m t y v c e e s t n r v

( i n n b i i c u s

l t g g i i

l e u d n

e e a t e n n d d p A p l r z s e n a h a a e n n i l t a t p a r e m h h a a s

a a

t u

a t t t d l C s C s S D V L A d p r i i o a p s m o e l e u y B H S R P

Galle Medical Journal, Vol 21: No. 1, March 2016 23 Reviews

Slow repetitive nerve stimulation Table 5 indicates mean (SD) score of individual Table 3 shows decrement response at rest, domains of MMSE in the patients and the controls. immediately after isometric exercise, two minutes The total score of MMSE, orientation and the after the exercise, post exercise facilitation and language domains of MMSE showed significant post exercise exhaustion. Statistically significant impairment in the first assessment of the patients difference of decrement response was observed at compared to the controls. There was no significant impairment when scores in the second assessment rest (A4) and two minutes after the exercise (C4, C5) in the second assessment compared to the controls. was compared with the controls (23). The decrement response was significant at rest

(A4, A5) and two minutes after the exercise (C4) in Discussion the second assessment compared to the first Small magnitude adverse difference of SNCV, assessment. MNCV, amplitude and area of CMAP on distal Seven occasions in the patients in the first stimulation and F-wave occurrence in acute OP assessment and five occasions in the controls showed poisoned patients compared to the controls was more than 8% decrement response either to the observed. Although it is a small magnitude, the second, fourth or fifth stimuli. difference is clinically significant as any damage to be evident by electro physiologically nearly 50% of the fibers need to be damaged as the remaining BAER unaffected fibers compensate the affected ones. The differences of the mean latency of I – III, III – V Therefore, the absolute values are greatly and I – V were prolonged in the test group compared underestimated. to the controls except in right side I – III difference The results of autonomic function tests showed in the first assessment of the test, right side I – III and significant impairment of heart rate variation during I - V difference in the first and the second assessment deep breathing and the patients had four fold residual in the test group compared to the controls. None of urine in their bladder compared to the healthy the differences were statistically significant (Table subjects. 4) (22). There was no detectable NMJ dysfunction assessed with exercise modified slow RNS or significant MMSE difference in the inter peak latencies of BAER after clinical recovery of acute OP poisoning. There were 60 patients and 61 controls. The mean age (SD) of the patients and controls were 31.5 (11.6) Although there was a slight transient cognitive years and 31.3 (11.8) years respectively. Forty-two impairment detected with the screening tests patients turned-up for the follow up assessment at following acute OP exposure, no long term cognitive six weeks following the exposure. Three out of 60 defects was detected by MMSE in clinical settings. patients required Intensive Care Unit (ICU) Diabetes is well known to cause neuropathy. admission. Therefore we excluded patients with diabetes from Seventeen out of 60 patients in the first assessment the study. Occupation matched controls were and 9/61 in the controls scored less than 24 in the recruited since there is evidence that occupational MMSE (X2 = 3.3, p = 0.06). Seven patients out of 42 exposure to pesticides can cause neuropathies. Very showed MMSE score less than 24 in the second few studies were found in the literature which assessment. There was no significant difference focused on SNCS / MNCS following acute ingestion between the number of patients with impaired of OP. There have been no studies which have looked MMSE in the second assessment vs controls (X2 = at the effects of acute OP exposure on peripheral 0.07, p = 0.7) and the second assessment vs the first nerve function compared to matched controls. Most assessment (X2 = 1.8, p = 0.17). All three patients large studies have examined nerve function in farm who had ICU admissions had MMSE score more workers who had chronic, probably low level than 26. exposure to pesticides.

24 Galle Medical Journal, Vol 21: No. 1, March 2016

Reviews

t n e

1 5 1

I 3 2 3 3 2 5 8 .

0 . . . . . m . . 0 3 . . 0 . C 0 1 1 1 0 s 0 - 0

0 - - 0

s o

o o o o o

t o t t t t t e o

% o o t t

s 9 t t

8 6 9 3 1

5 s 9 . . . . . 4 0

5 . . . 4 5 a 9 1 2 2 2 - 0 - 3 - - 2 - - - 0 - - - - d n 2

s v

t n ) t e e

d n

e n

n s 2 a 2 2 e 7 3 9 2 7 8 5 0 4 s m ...... r – e 0 0 s e . . 0 2 1 1 0 0 0 3 2 e t s s ------0 0 f s M s e f 1 i s a ( s d t s a 1

8 8 7 7 t 5 8 3 9 . . I 5 0 0 . . . . 3

. . n 2 2 - 5

0 1 2 - 1 - - -

C 0 0

e - - - -

- -

o t o o t o

o t o o o m

t t

% o o

t t t t

8 s t t

2 . 5 5 . 1 2 s 2 6 5 1 . 0 - 0 1 2 . . . . 0 9 . . . e - 0 1 0 0 1 s - 0 0 0 s s l a o d r n t 2

n s e o – v

)

s c s s l

t n l n

o e n 3 4 e 5 3 6 9 5 4 9 1 a o

4 r ...... r 0 0 . e r e

t h . . 1 1 0 0 1 0 2 3 i t e 0 ------t t n 0 0 f n

M f a o i o d p c d ( C n a

s t n

6 e I

2 7 1 i 0 . . . 1 2 1 2 t 4 9 4 . . . . t - - - - 0 1 0 C n 0

- 0 0 - 1 - a -

- - -

e o o o o

t t t t o

o o p

% t o t t o o o

t t 6 9 2 t 8 5

. . . . s 9 2 1 e

. 2 . . 9 2 2 1 0 1 1 0

s . 1 1 0 h e 0 t s

s f a o t

e s 1 c n

n s o e – i v )

t s s s e t l s l f n o 3 6 2 3 7 2 7 f u o 9 9 6 3 . . . . r i 0 0 0 . . . . e r t . . . 0 0 0 0 i a t 0 0 0 0 d

- - - - t 0 0 0 n

n h

o n

o p x c a

( e C e

e M s i c r

e t

n x ) ) ) ) ) e ) ) ) ) ) ) e % 4 3 3 2 7

5 5 5 6 0 0 ) . . . 0 0 ...... t m d . . 0 0 0 n

0 0 0 0 1 1 n E s ( ( ( s 0 0 ( ( ( ( ( (

a ( ( s

2 S

o 9 3 2 e 8 8 4 6 6 5 . . . e (

0 0 ...... s p 0 0 1 0 0 0 0 1 1 - - -

1 1 s M

s a d t n n e a i

t t a n

P o e i ) ) ) ) ) ) ) ) ) ) ) t % 3 5 m 5 5 5 5 5 5 4 4 5

) ...... a 0 0 s . . t 0 0 0 0 0 0 0 0 0 n s E ( ( ( ( ( ( ( ( ( i 0 0

a e l ( ( S

4 5 0 2 2 2 6 2 8 s i e ...... (

9 0 s c . .

1 1 1 1 0 0 1 1 0

a ------0 1 M a t f s

1 e s i ) c r E e ) ) s ) S ) ) ) ) ) ) ) ) l 4 4 ( 5 x 3 4 5 5 5 5 4 5

o 0 0 ...... e r 0 0 0 0 0 0 0 0 0

t 0 0 % ( ( ( ( ( ( ( ( ( ( ( t

s 5 6 5 9 5 8 6 3 6 n 5 1 ...... o 0 0 o a 0 0 0 0 0 0 1 1 1

. . C ------e 1 1 p

, M e s n o

) ) )

p 2 4 5 s ) )

1 1 e

B B B

l l l ( ( ( r r r r A A

a a a

e e / / e e e e t t t 1 1 t f f s s s m m m f i i i i i i a a a n

c c c

x x x C B ) ) ) r r r y y ( ( e a a a y 2 4 5 r

l l e e e l e e e n n e x x x m m m t t t t

) ) ) C C C m e e e o o r r r 2 4 5 a a ( ( ( a

e i i i i

i e t t e e e c c c t t t e e e s d d i i i a d r f f f A A A m t r r r s s s e e u e ( ( ( i t t t i i i a a a c a

l a

e e e c c c t t t i r m m m e r r r h n n n s s s c i i i a m m m e e e x e e e m m a m r r r i i i o o o x x x f e D P m m m

s s s

e e e t t t t t t i i i

e e

a a a a a 2 2 2

a s s c c c

l l l

: i i i i i t t t t t t t t t a a a r r r

c c 3 n n n n n n n n n t t t r r

e e e e e e e e m m m e e e e e e i i i e x x m m m m m m m m x x x m l m m m e e e e e e e e e e a a a e o o o

r r r r r r r r r b t t s s s c c c c c c c c m m m i i i c s s a e e e e e e e e e o o T D D D D D D D D D P P

Galle Medical Journal, Vol 21: No. 1, March 2016 25

Reviews

t t s n t n v 4

4 6 e n 7 5 7 e 4 2 2 t . . 7 1 0 . . . e 1 . 0 0 0

n . 0 0

. 0 . . . m 0 0 0 m 0 . e - - m 0

0 0 0 s

s s o

m o o o s s s o o o o o o t o t t t s e o t

t t t t t e e v t

s 7 s

7 3 5 e 2 8 9 5

s s 4 5 s . . . 1 . 1 0 s . .

. . 0 0 1 .

a a

0 . . . s

0 0 0

0 0 2 1 -

0 d - - - a t

0 0 0 d - - - - n -

- - - n t

s 2

1 2

9 t

4 3

6 4 . 0 n

t 0 . . . I

. 0 e

– n 1 0 0 6 0 -

s 2 1 5 6 5 C

- . . 0

I s m v

0 0 0 . o o o

l o 0 0 . . . s m o t t t C

0 o s t

o 0 0 0

s % s l

t o o

r s

o t t 1 9 2 e o t 6 o o o

t % . . 8 e r 5 -

s t t t

. 7

. t s 5 8 n

5 . 0 0

s 9 8 0 s n

0 1 1 0 2 0 - - 9 o 0

a -

. . . . . 0 a -

. -

0 0 0 0 0 C d

0 C - - - - - n

1 t

3 1

4 0 . . n

2 s . .

3 . 0 0 e

. v t 0 0

- - 0

0 n 9 - s m s

7 1 1 1 1

o o e 0 . . . . o o l s v t t 0 . o t t

. 0 0 0 0 s o t

m 0

s t

r 0 7 2 l e s

o o o o

5 9 t .

s o 1 t t t t s . . o 0

1 o t

- n r

. 0 e s

t - 2 0

t 4 3 9 9 - s

o a 0 4 - -

n s 0 0 2 0 0 -

0 . . . . . a

o C .

0 0 0 0 0 t

0 C s - - - - -

n e

m Table 5: Score of individual domains in the MMSE s m s 4 v t 4 0 1 1 1

s s

t 0 . . . . . n

s . e n e 1 1 0 0 0 e s 0 e - - - - -

s s - m 5 4 5 4 2 3 s a m s 0 0 0 0 0 0 s s a

. . . . . 0 d s e . st nd t n 0 0 0 0 0

e s s 0

2 s

MMSE / Domain Controls 1 2 Mean difference 95 % CI s 1

s a

e a

c t st nd st st nd st n assessment assessment d s

s (n = 61)

l n n

Controls - 1 Controls - 2 1 assessment - Controls vs 1 Controls – 2 1 assessment t 1

2 o n nd r

r s e -

(n = 60) (n = 42) e

assessment assessment 2 assessment assessment assessment vs t e

5 f s

4 1 c m f t n 3 1 4

l . . 0 i nd . . . s n . n o o

0 0 s e d e 0 0 0

2 assessment 0 r - -

c r v e t -

e m n s s f n 7 e s l 8 3 3 3 s a f 1 s 0 i o o h 1 0 0 0 0 a e

e r . . . . . 0 MMSE total 25.1 (3.3) 26.5 (2.8) 26.6 (3.3) -1.4 -0.05 -1.4 -2.5 to -0.3 -1 to 1 -2.4 to -0.4 d t . t

s C

0 0 0 0 0

s 0 n n M d a a o score e d n t C

n s

t a 2 M

1 n

s -

Orientation 9.4 (0.9) 8.8 (1.4) 9.8 (0.5) 0.6 -0.4 -1.0 e -1 to -0.2 -0.7 to -0.09 -1.5 to -0.6 t

s 4 1 n m 6 4 5 l . . t . . . s

e 0 o n 1 0 i s s 0 0 0 r - - e t

Registration 2.9 (0.2) 2.9 (0.3) 2.6 (0.6) 0 0.3 -0.04 -0.07 to 0.07 -0.6 to -0.3 -0.1 to 0.04 e v t

s a m n s s l s 7 2 1 3 3 6 p o s o a

0 0 0 0 0 0 r

...... e C e t

Attention and 3.2 (1.8) 2.8 (1.8) 3.0 (2.0) 0.4 0.1 -0.1 -1 to 0.3 -0.9 to 0.6 -0.7 to 0.4 s 0 0 0 0 0 0

n s h

a o t

t calculation t C s

) n n ) ) ) ) )

) 1 3 i e . 5 6 0 6 3 2 . . . . . E

3 4 d m 0 0 2 0 1 R (

n Recall 2.4 (0.8) 2.6 (0.8) 2.6 (0.6) -0.1 -0.1 -0.1 -0.2 to 0.4 -0.2 to 0.4 -0.3 to 0.2 S s ( ( ( ( (

= s 2

E n 6

. 8 6 0 6 2 e M e

. . . . . n A s 2 5 2 2 2 2 6 ( ...... 9 2 3 2 8 m s 2 3 Language 8.5 (0.8) 8.0 (1.4) 8.2 (1.3) 0.5 0.4 -0.1 -0.9 to -0.1 -0.8 to 0.04 -0.5 to 0.2 M s 0 0 0 0 0 0 B a

5 s

e f ± ± ± ± ± ± e

h o

1 8 9 1 8 9 s

N t ......

a e 2 1 3 2 1 3 ) p n ) ) ) ) ) d n c

) 8 u n i e . 4 3 8 8 4

0 n o 2 . . . . . s r 2 t 6 e m 1 0 1 0 1 ( s

g n s r ( ( ( ( (

1 t = s e 5

s t a 8 9 8 6 0 . e f . . . . . e

n n f s 6 ( e i T 8 2 2 2 8 m s

2 2 2 2 2 2 2 ...... a o d m 0

0 0 0 0 0 0 s

y ± ± ± ± ± ± l e =

c ) s a ) ) ) ) ) ) 1 9 0 1 9 7 s N s n ...... 3 l u 1 9 2 8 8 8 .

a e 2 1 4 2 1 3 . . . . . o t 6 t 3 d s r

0 0 1 0 0 i ( t a 1 ( ( ( ( (

v = l

i 1

4 9 2 4 5 . o n

. . . . . d

n 5 (

9 2 3 2 8 C

n 6 2 3 2 2 2 a l 2 ...... i o e

p 0 0 0 0 0 0 0 r f

u 7 t ± ± ± ± ± ± o m o n =

r n o

2 8 0 1 8 9 i e N e g

......

d C a r

2 1 4 2 1 3 h l

n o n a

m n a T c t o n

o i o e o

S o t n i t : i

g t

D a

t o

r a a : i 4 e

/ a E

t t

t l d u 5 l e s i S n e

n e

i u g a E d s l

r e e i e

c g c i n l t t V V S s 1 1 M l b o

r t - - e e a h 1 1 t b a c a f 1 1 g 1 V 1 V M M s O R A c R L a i

e - - - - 1 1 T

T R 1 1 1 L 1 1 1 M

26 Galle Medical Journal, Vol 21: No. 1, March 2016 Reviews

Reduction of SNCV and MNCV of the patients in our respectively. Improper nursing care may be partially study indicate that there may be demyelination contributed to the nerve damage. following acute OP exposure. Since we did not Although autonomic function in farm workers has follow up the patients beyond six weeks of exposure been looked in to, there were no studies that looked we do not know whether the changes were reversed at autonomic function following acute OP exposure. or worsened with time. In the current study neurophysiological parameters Reduced amplitude and/or area of CMAP on distal of the exposed individuals were compared with the stimulation were observed in several comparisons. parameters of matched controls. These indicate that there may be an axonal damage Variation of heart rate during rest, deep breathing since amplitude and area under negative curve and isometric muscle contraction were tested in of CMAP are directly proportional to the number flower bulb farmers who had chronic mixed of functioning axons (10). If the whole length of pesticide exposure by Ruijten M.W.M.M et al. the nerve is affected, F-wave latency should be (1994). Significant autonomic dysfunction was prolonged. Reduced nerve conduction velocity only shown in variation of heart rate during rest and deep in the distal segment may be an evidence of distal breathing (24). We did not assess variation of heart demyelination with sparing of proximal segment. rate during rest but results for heart rate variation Since we did not perform segmental nerve during deep breathing were similar in both studies. conduction studies focal damage cannot be excluded. In the tests reflecting parasympathetic damage (heart rate response to Valsalva manoeuver, heart F-wave studies are also important for recognizing the rate variation during deep breathing and immediate proximal segment involvement since it is produced heart rate response to standing) a positive result by antidromic activation of motor neurons. It is also was shown only with heart rate response to deep important to identify marginal changes of standard breathing. It is well known that the response of MNCS since an impulse travels a long pathway to the heart rate can be abolished by atropine (7). The produce an F-wave, and additive effects are more first assessment of the patients was conducted obvious in F-wave latency measurement as it within the mean of three (IQR 1 - 4) days after the measures the delay in a much longer segment (10). cessation of atropine therapy. The effects of atropine The latency of and F-wave includes the time may not have disappeared completely even 6 weeks required for the evoked action potential to ascend after exposure. On the other hand, only 48/66 antidromically to the anterior horn cells, the time patients were able to do the Valsalva manoeuver. between the beginning of antidromic activation and The sample size may not have been adequate to subsequent orthodromic discharge (the central draw a conclusion. Or else the autonomic function “turnaround” time) and the time required for the of exposed individuals may not have been affected. resultant action potential to descend orthodromically from the anterior horn cells to the muscle fibers. Change of diastolic BP 3min after standing was F-wave abnormalities may be loss of response significantly low in the patients, but recovered (reduction of occurrence) or prolonged latencies completely at six weeks after exposure. Blood (10). F-waves do not occur with each stimulation. pressure response to standing reflects sympathetic They depend on excitability of motor neurons. function. But it begins to show abnormal results In normal individuals F-wave occurrence is more with more severe sympathetic nerve damage (7). than 90% (10). Sympathetic damage in our patients may not have been severe enough to give abnormal results in the There was a statistically significant reduction of blood pressure response to standing. F-wave occurrence observed in the median and the ulnar nerves in the first assessment compared to Reduction of amplitude and prolongation of latency the controls. Although there was prolongation of in SSR are in favor of sympathetic damage at one F-wave latency, it was not statistically significant. week after exposure. SSR represents the change Some nerves such as the ulnar and the common in voltage measured at the skin surface following peroneal are highly vulnerable to external damage in a single electrical stimulus. It depends on the the ulnar groove and behind the head of the fibula electrical activity arising from sweat glands (10).

Galle Medical Journal, Vol 21: No. 1, March 2016 27 Reviews

Latency of SSR is inversely proportional to the A study done to see the effects of occupational number of sweat glands innervated (10). Hence exposure to OP on neuromuscular function did not sympathetic innervations to sweat glands were less show any detectable neuromuscular disturbances in the patients compared to the controls. In the with RNS and with SF-EMG (28). second assessment both the latency and the The frequency of neuromuscular transmission amplitude of SSR were improved. We could not abnormalities detected by SF-EMG was eliminate the effects of atropine on sweat glands significantly higher than those detected by RNS and changes in the first assessment may have been (28, 29). due to effects of atropine. However patients did not show sympathetic dysfunction as reflected in Decrement response is more obvious in fast SSR at six weeks after exposure. repetitive stimulation than the slow repetitive stimulation. The current study concentrated only It is well known that high residual urine volume is the slow repetitive stimulation with exercise associated with increased risk of urinary tract modification to magnify the function of NMJ. infections. Since our results showed that residual However this may not be sensitive enough to detect urine volumes in patients were significantly higher subtle abnormalities (30). than in the controls, it is very important to follow up such patients further. The amplitude of compound muscle action potential may increase during high-frequency nerve One third of patients were admitted with co- stimulation or after voluntary activation of muscle ingestion of alcohol. Acute alcohol ingestion is (10). The measurement of CMAP amplitude in unlikely to have had any effects on the parameters we fast repetitive stimulation is not recommended by assessed, since the assessments were carried out at some since amplitude of the CMAP during high one week after the exposure. It is well known that frequency stimulation may give variable and long term alcohol consumption is associated with inconsistent results (15). Therefore we measured neuropathies including autonomic neuropathy. CMAP amplitude just after and two minutes after the Adjusted multiple linear regression did not show isometric muscle exercise. Even with isometric widespread effects of alcohol consumption on muscle exercise the current study was not able to autonomic function. demonstrate significant decrement comparing to the The number of patients who could complete the test controls. Previous studies on neuromuscular synapse of blood pressure response to sustained hand grip function in organophosphate workers also did not was significantly lower than the number of controls show significant decrement response compared to who could complete the test. The difference may be the controls (13,14). due to muscle weakness following poisoning. With each depolarization, Ca++ is release into the The gold standard electrophysiological tool to periterminal space. If another depolarization occurs, explore the neuromuscular junction function is high Ca++ concentration leads to increase in single-fiber electro myography (SF-EMG) (15, 25, acetylcholine quanta released. If neuromuscular 26). We explored whether RNS with exercise might transmission is impaired this greater acetylcholine be a useful alternative method as SF-EMG is not release briefly improves synaptic transmission, available in most centers. producing facilitation (15). Marked facilitation is Few patients had RNS abnormalities in the first characteristic of presynaptic blockade. Although assessment and this was no different from the facilitation can also sometimes be positive with controls. Results of RNS six weeks after the postsynaptic problems (15). exposure showed significant decrement response. Although it was recommended to calculate the However RNS does not appear to be a useful method changes in CMAP size between the first and the to explore medium to long term NMJ function. fourth or the fifth response of a train of stimuli (15) NMJ dysfunction has been demonstrated by fast we took the second response as well, since Maselli repetitive nerve stimulation with OP poisoning et al (2004) and Jayawardana et al (2008) observed before clinical recovery (27), but there is no evidence the maximum decrement at the second stimulus whether these changes persist over the long term. (27,31). The data set of the current study showed

28 Galle Medical Journal, Vol 21: No. 1, March 2016 Reviews

the maximum decrement either at the second or the Cognitive function was tested with the simple and third stimulus. quick assessment tool of MMSE. The MMSE The majority of the cases of the current study provides a measure of cognition which covers a showed increment pattern progressively over the broad set of cognitive domains: orientation, train. For that reason the mean percentage registration, short term memory, attention, decrements shown in the table 2 are negative values. calculation, visuo-spatial skills and apraxia (18). This pattern is described as pseudo increment and is The sensitivity and specificity of MMSE were 100% a normal variant (32). and 69% to the cut off level of less than 24 evaluated against the performances at CAMCOG (Cambridge Even though a decrement response (>8%) was Cognitive Score) (18). observed in few occasions both in the patients and the controls, even these decrements were not Stephens R et al (1995) reported impaired cognitive confirmed. False positive results of repetitive nerve function in sheep farmers who were exposed to OP stimulation is possible since the studies are through sheep dipping, which involves immersing technically difficult; artifacts resulting from each animal in a pesticide solution to control electrode placement, movements, changes of skin parasitic infections. Exposure to OP was appeared resistance and intramuscular temperature cannot be to be splashing on to the skin where protective entirely controlled (25). cloths were seldom worn. The speed of performance of simple reaction time, symbol-digit substitution Very few human studies have examined ototoxic and syntactic reasoning were significantly slower in effects with pesticide exposure. No studies have the farmers than the controls. No impairment was assessed the ototoxic effects following acute found in short term memory (by digit span test and exposure to OP. visual spatial memory) and long term memory (by BAER is a sensory evoked potential used to assess serial word learning, category search classification, the integrity of the auditory pathway from the eighth category search recognitions) in the cases compared nerve to the auditory cortex (16). The inter peak to the controls (1,37). latencies (I - III, III - V and I - V) of BAER provide a Steenland K et al (1994) investigated chronic squeal straightforward method to roughly localize lesions of acute occupational OP poisoning (38,39). They in the auditory pathway (16). studied cognitive function via finger tapping (motor Animal studies by Bielefeld E C et al. (2005) and speed test), sustained visual attention, visuomotor Harris K C et al. (2006) showed shifting of evoked accuracy, visuomotor speed, symbol digit, visual potential threshold and loss of inner and outer hair memory and serial digit learning tests. Significant cells following application of 10mM, 5mM and impairment found in sustained visual attention and 3mM of PQ to cochlea through a surgical incision symbol digit tests in the cases compared to non- (33,34). The current study did not look into the exposed reference (38,39). auditory threshold level. A study done on volunteers with varying degrees of Clerici W J (1996) demonstrated disruption of occupational exposure to OP showed significant generation and transmission of afferent cochlear impairment on the Bender Visual Motor Gestalt test signals by generation of reactive oxygen species (used to evaluate visual motor maturity) and part B within the perilymphatic space with artificial of the Trail Making test (Trail Making test is perilymph and hydrogen peroxide (35). Even neuropsychological test of visual attention (37). though OP produces reactive oxygen species after ingestion, the concentration of substances reaching The study done by Fiedler N et al (1997) recruiting the perilymphatic space may not be sufficient to tree fruit farmers who had occupational exposure to damage the cochlea. OP demonstrated significantly slow simple reaction time for the dominant and nondominent hands of Kimura K et al. (2005) studied the effects of the the exposed than the controls. No significant event-related evoked potentials in tobacco farmers impairment seen in the visuomotor coordination, who were exposed to different pesticides including verbal memory, verbal ability and expressive and OP. They did not see any significant effects on the receptive language (38,39). event-related evoked potentials (36).

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The review article by Kamel F et al. (2004) 4. Annual Health Bulletin. Morbidity and Mortality. 2003, looked into association of pesticide exposure Health Ministry Sri Lanka. 21-30. with neurological dysfunction. They found 5. Singh S and Sharma N. Neurological syndromes following inconsistencies among the studies. Most studies organophosphate poisoning. Neurol India, 2000; 48(4): 308- found deficits in one or more tests of cognitive 13. function but different tests were affected in different studies (40). 6. Ewing DJ, Testing for autonomic neuropathy. Lancet, 1981; 1(8213): 224. The current study did not showed persistent impairment of cognitive function following acute 7. Ewing DJ, et al. Cardiac autonomic neuropathy in diabetes: OP exposure. The methods we used may not be comparison of measures of R-R interval variation. sensitive enough to detect mild cognitive Diabetologia, 1981; 21(1): 18-24. impairment. Further studies are required with more 8. Hilz MJ and Dutsch M. Quantitative studies of autonomic sophisticated neurobehavioural tests to assess function. Muscle Nerve, 2006; 33(1): 6-20. cognitive function following acute OP poisoning. 9. Ewing DJ and Clarke BF. Diagnosis and management of diabetic autonomic neuropathy. Br Med J (Clin Res Ed), Acknowledgement 1982; 285(6346): 916-8. The studies were funded by the South Asian Clinical 10 Gerawarapong C. Association of peripheral autonomic Toxicology Research Collaboration, which was neuropathy and sympathetic skin response in the patients funded through the Welcome Trust / NHMRC with diabetic polyneuropathy: A pilot study in Thailand, Australia. I would like to acknowledge participants, 2015 Dec; 98(12): 1222-30. the consultants who gave their patients for the 11. Kelly CE. Evaluation of voiding dysfunction and study, consultant radiologists and the staff under measurement of bladder volume. Rev Urol, 2004; 6(Suppl them, administrative staff and health care 1): S32-7. professionals at the Teaching Hospital, Karapitiya 12. Guidelines EBM. Determine the volume of residual urine by and the General Hospital, Matara, Heads and the staff ultrasonography. EBM Guidelines, 2004. members of the Departments of Pharmacology, Medicine, Parasitology, Microbiology and 13. Misra UK, et al. A study of nerve conduction velocity, late Pathology, Faculty of Medicine, University of responses and neuromuscular synapse functions in Ruhuna, the members and the Clinical Research organophosphate workers in India. Arch Toxicol, 1988; Assistance of the South Asian Clinical Toxicology 61(6): 496-500. Research Collaboration. A special word of thank 14. Engel LS, et al. Neurophysiological function in farm is extended to Professor A H Dawson, Professor workers exposed to organophosphate pesticides. Arch N Buckley, Professor A Fernando, Professor K D Environ Health, 1998; 53(1): 7-14. Pathirana, Professor P L Ariyananda and Professor th B Perera. 15. Aminoff MJ. Electrodiagnosis in clinical Neurology. 5 ed Eelectromyography, nerve conduction studies, and related techniques, ed. A. M.J. Vol. 1. 2005, USA: Elsevier. 231- References 425.

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