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Effects of drugs on the formation of homovanillic acid in the rat retina
European Journal o f Pharmacology, 40 ( 1 9 7 6 ) 1 7 5 - - 1 7 8
175
© E l s e v i e r / N o r t h - H o l l a n d B i o m e d i c a l Press
Short communication EFFECTS OF D R U G S ON THE F O R M A T I O N OF HOMOVANILLIC ACID IN THE RAT RETINA BEN H.C. W E S T E R I N K * a n d J A K O B K O R F **
* Laboratory for Pharmaceutical and Analytical Chemistry, Department o f Clinical Chemistry, Antonius Deusinglaan 2, and ** Department o f Biological Psychiatry, State University, Groningen, The Netherlands Received 6 A u g u s t 1976, a c c e p t e d 9 A u g u s t 1 9 7 6
B.H.C. W E S T E R I N K a n d J. K O R F , Effects o f drug on the formation o f homovanillic acid in the rat retina, E u r o p e a n J. P h a r m a c o l . 40 ( 1 9 7 6 ) 1 7 5 - - 1 7 8 . H o m o v a n i l l i c acid ( H V A ) levels were m e a s u r e d in t h e eye a n d t h e c o r p u s s t r i a t u m o f rats u n d e r n o r m a l condit i o n s a n d a f t e r d i f f e r e n t drug t r e a t m e n t s . N e u r o l e p t i c agents such as clozapine, c i s - f l u p e n t h i x o l a n d h a l o p e r i d o l i n d u c e d c o m p a r a b l e increases in H V A levels, whereas t h e n o n - n e u r o l e p t i c t r a n s - i s o m e r of f l u p e n t h i x o l was inactive in b o t h s t r u c t u r e s . A p o m o r p h i n e decreased H V A levels in t h e r e t i n a a n d t h e c o r p u s s t r i a t u m , w h i l e amp h e t a m i n e i n d u c e d a decreased H V A f o r m a t i o n in t h e r e t i n a a n d did n o t c h a n g e levels o f H V A in t h e c o r p u s s t r i a t u m . P r o b e n e c i d caused a similar p e r c e n t a g e rise of H V A in b o t h s t r u c t u r e s . M o r p h i n e a n d o x o t r e m o r i n e i n d u c e d a rise in H V A levels in the c o r p u s s t r i a t u m b u t n o t in retinal samples. H o m o v a n i l l i c acid Corpus striatum
Oxotremorine
Morphine
1. Introduction In addition to the dopamine (DA)-containing nigrostriatal and mesolimbic pathways, dopaminergic processes have been described in the retina. The retinal DA-containing neurons have been histochemically characterized and were found to be similar in morphology to amacrine cells (e.g. H~iggendal and Malmfors, 1965; Ehinger, 1966). Many centrally acting drugs e.g. neuroleptics, central stimulants, analgesics and cholinomimetics are known to influence the formation and utilization (the turnover) of DA in the central nervous system. Measurement of the levels of the two main DA metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) appeared to be a very sensitive m e t h o d to detect changes in the turnover of DA; there have thus been many studies of the formation of these metabolites in brain during various drug treatments (e.g.
Neuroleptics
Retina
Laverty and Sharman, 1965; Westerink and Korf, 1976). From studies on the formation of DOPAC and HVA in various regions of the rat brain it appeared that in general striatal and mesolimbic structures respond in a similar manner to drug treatment (Westerink and Korf, 1976). Apparently these cerebral dopaminergic systems have a comparable vulnerability to various drugs. The presence of DA in the retina however allows one to study the influence of drugs on DA metabolism in a neuronal environment which differs from the striatal and mesolimbic structures. A recently developed sensitive assay for HVA (Westerink and Korf, 1976) made it possible to investigate the formation of HVA in a single rat eye. We studied the influence of amphetamine, apomorphine, clozapine, cis-flupenthixol, trans-flupenthixol, haloperidol, morphine, oxotremorine, pargyline and probenecid on the formation of HVA in the retina. The effects of these drugs on re-
176
tinal and striatal DA-metabolism will be compared.
B.H.C. W E S T E R I N K , J. K O R F
mental values were analyzed with the Student's t-test. A p value of less than 0.05 was considered significant.
2. Materials and methods 3. Results Male albino Wistar rats of 1 7 5 - - 2 5 0 g weight (T.N.O., Zeist, The Netherlands) were used. The following drugs were used: amphetamine sulphate, apomorphine hydrochloride, clozapine, cis-flupenthixol hydrochloride, trans-flupenthixol hydrochloride, haloperidol, morphine hydrochloride, oxotremorine sesquioxalate, pargyline hydrochloride, probenecid. The doses of drugs given in Results are expressed as free base or acid. Apomorphine was dissolved in saline containing 25 mg ascorbic acid/100 ml. Haloperidol was used as a commercially prepared solution (Serenase®). Clozapine was dissolved in a drop of acetic acid and diluted with saline. All other drugs were dissolved in saline. Groups of drug-treated rats were matched with 3--4 animals injected with saline, which were considered as controls. Drugs were administered intraperitoneally and the animals were killed following the time schedules given in Results. Because HVA was found in both retinal tissue and the aqueous humor, we analyzed total eyes and considered these samples as "retinal samples". The eyes were taken out, frozen on dry ice and homogenized separately in 1.0 ml 0.4 M perchloric acid. HVA was assayed according to our previously described method (Westerink and Korf, 1976). The method is based on isolation of HVA on small Sephadex G 10 columns, followed by automated (continuous flow) fluorometric detection. Recoveries for 25 ng HVA added to cerebellar tissue were between 80 and 90%, and no corrections were made for these recoveries. Levels given for retinal as well as striatal samples are the mean of the bilateral structures, which were analyzed separately. Difficulties in obtaining correct tissue blanks prevented the determination of DOPAC (Westerink and Korf, 1976). Differences between control and experi-
Mean HVA values of saline-treated animals were (±S.E.M.) in the retinal samples: 4.3 ± 0.2 ng/eye (n = 12), and in the corpus striarum: 0.45 + 0.01 pg/g (n = 20). As after pargyline treatment (75 mg/kg, 2 h), the fluorometric assay gives values below the detection limit (1.5 ng/sample) in both striatal and retinal samples, the fluorescence recorded can be completely attributed to the presence of HVA. Table 1 shows the effects of drug treatment on HVA formation in the retina and the corpus striatum. There was significant decrease in HVA levels in both structures after apomorphine treatment (5 mg/kg, 45 min), while amphetamine (5 mg/kg, 45 min) only induced a decrease in HVA in the retinal samples. The neuroleptic agents clozapine (15 mg/kg, 2 h), cis-flupenthixol (2 mg/kg, 2 h) and haloperidol (1.0 mg/kg, 2 h) induced a significant increase of HVA levels in both retinal and striatal samples, while trans-flupenthixol (2 mg/ kg, 2 h) did not change HVA levels in either sample. After injection of morphine (20 mg/kg, 2 h) and oxotremorine (1.0 mg/kg, 60 min) significant increases of HVA levels were found only in the corpus striatum. The morphine and oxotremorine treatments were repeated in combination with probenecid (200 mg/kg, 75 min). Again no changes in HVA levels were observed in the retinal samples, although there were pronounced increases of HVA levels in striatal samples.
4. Discussion
The formation and utilization of DA in the brain appears to be very sensitive to centrally acting drugs. Many drugs, including neurolep-
H V A IN THE R E T I N A
177
TABLE 1 E f f e c t s o f various t y p e s o f drugs on H V A levels (2 S.E.M., n) in retinal samples (total eyes) and c o r p u s s t r i a t u m o f t h e rat. Drug
Dose (mg/kg)
Killed after injection (rain)
Retina (ng/eye)
Saline Amphetamine Apomorphine Clozapine Cis-flupenthixol Trans-flupenthixol Haloperidol Morphine Oxotremorine Probenecid
-5.0 5.0 15 2.0 2.0 1.0 20 1.0 200
60 45 45 120 120 120 120 120 60 120
Probenecid controls
200
75
8.0 ± 0.6 ( 7 )
1.25 -± 0.05
7)
Probenecid Morphine
200 20
75
6.7 ± 0.6 ( 5 )
2.40 ÷ 0.18
5) c
Probenecid Oxotremorine
200 1.0
75
7.7 ± 0.6 ( 6 )
1.92 ± 0.08
6) c
4.3 2.8 2.7 5.2 11.0 4.4 9.9 4.9 4.6 8.8
± 0.2 _+ 0.4 ± 0.3 ± 0.4 ± 0.7 ± 0.3 ± 0.5 ± 0.6 ± 0.3 -+ 0.6
(12) (5) (10) (9) (3) (3) (4) (8) (9) (4)
b c a c c
c
S t r i a t u m (pg/g)
0.45 0.48 0.17 1.24 2.57 0.45 2.97 1.09 0.90 1.03
± 0.01 ± 0.03 ± 0.03 ± 0.08 -± 0.18 ± 0.03 ± 0.27 ± 0.04 ± 0.09 ± 0.7
(20) (10) (5) c (7) c (3) c (3) (9) c (12) c (6) c (12)c
D i f f e r e n t f r o m c o n t r o l : Cp < 0.001; b p < 0.005; ap < 0.02.
tics (e.g. Laverty and Sharman, 1965), apomorphine (Roos, 1969), amphetamine and anorectics (Jori and Dolfini, 1974), cholino-. mimetics (Laverty and Sharman, 1965), analgesics (Sharman, 1966), LSD (Da Prada et al., 1975), antidepressants and anti-epileptics (Westerink et al., in preparation), were found to influence DA metabolism. The various mechanisms by which these drugs interact with DA turnover need further investigation. The presence of DA in the retina allows the drug-induced changes in HVA formation in the forebrain to be compared with those in a differently organized neuronal structure. We suggest that a drug acting directly on dopaminergic neurons as a result of its antagonistic or agonistic behavior at dopamine receptors, does so independently of the neuronal environment. However, an indirectly acting drug could give different responses in differently organized structures. There is considerable evidence that neuroleptics act as antagonist and apomorphine as an agonist at dopamine receptors in the brain.
Neuroleptic treatment results in an increased DA turnover in the brain and a rise in HVA levels, while apomorphine treatment causes a decreased DA turnover and decreased HVA levels (Roos, 1969). Inhibition of DA-sensitive adenylate cyclase by neuroleptics is thought to reflect their postsynaptic receptor blockade, while reversion of apomorphineinhibited synaptosomal tyrosine hydroxylase should represent their presynaptic receptor blockade (Iversen et al., 1976). These neurochemical changes induced by neuroleptics and apomorphine can be studied in striatal homogenates and are thus independent of the neuronal organization. It is therefore not surprising that neuroleptics increased HVA levels, while apomorphine reduced HVA levels in both retina and corpus striatum (table 1). The three different types of neuroleptics tested: clozapine, cis-flupenthixol and haloperidol induced comparable increases in HVA levels, whereas the non-neuroleptic transisomer of flupenthixol was inactive in both structtLres. Recently Da Prada et al. (1975) described the
178
agonistic behavior of LSD with respect to DA. The decrease of HVA levels found by these authors in both corpus striatum and retina again suggests a similar metabolic response to direct DA receptor stimulation in both structures. The decreased HVA levels in retinal samples after amphetamine treatment were not seen in corpus striatum, but the biphasic effect of this drug on DA metabolism with regard to both time and dose (e.g. Koe, 1975) could be responsible for this discrepancy. Probenecid pretreatment caused a similar percentage rise in HVA in both structures, which suggest an identical interaction with the HVA efflux in corpus striatum and retina. Morphine and oxotremorine did not induce a rise in HVA levels in the retinal samples as was seen in the corpus striatum. As combined treatment with morphine or oxotremorine with probenecid failed to elicit an additional accumulation of HVA in the retina, it seems unlikely that morphine and oxotremorine work similarly in both systems but with a different onset and duration. Morphine and oxotremorine caused identical changes of DOPAC and HVA levels in the corpus striatum (Westerink and Korf, 1976), thus we do not expect that DOPAC levels changed in retinal samples. If morphine and oxotremorine affect DA metabolism by indirect mechanisms dependent on the neuronal organisation of the corpus striatum and substantia nigra, it is conceivable that such a response is absent from retinal tissue. The results of the present study suggest that, in corpus striatum and regina, comparison of DA metabolism based on HVA levels could differentiate between drugs whose action is dependent on or independent of the connections of dopaminergic neurons with other neuronal systems.
B.H.C. WESTERINK, J. KORF
Acknowledgement The authors wish to thank Mr. Theo Mulder for expert technical assistance.
References Da Prada, M., A. Saner, W.P. Burkard, G. Bartholini and A. Pletscher, 1975, Lysergic acid diethylamide: evidence for stimulation of cerebral dopamine receptors, Brain Res. 95, 67. Ehinger, B., 1966, Distribution of adrenergic nerves in the eye and some related structures in the cat~ Acta Physiol. Scand. 66, 123. H//ggendal, J. and T. Malmfors, 1965, Identification and cellular localization of the catecholamines in the retina and the choroid of the rabbit, Acta Physiol. Scand. 64, 58. Iversen, L.L., M.A. Rogawski and R.J. Miller, 1976, Comparison of the effects of neuroleptic drugs on pre- and postsynaptic dopaminergic mechanisms in the rat striatum, Mol. Pharmacol. 12,251. Jori, A. and E. Dolfini, 1974, On the effect of anorectic drugs on striatum homovanillic acid in rats, Pharmacol. Res. Commun. 6, 175. Koe, B.K., 1974, Effect of neuroleptic drugs on brain catecholamines, in: Neuroleptics, eds. S. Fielding and H. Lal (Futura, New York). Laverty, R. and D.F. Sharman, 1965, Modification by drugs of the metabolism of 3,4-dihydroxyphenylethylamine, noradrenaline and 5-hydroxytryptamine in the brain, Brit. J. Pharmacol. 24, 759. Sharman, D.F., 1966, Changes in the metabolism of 3,4-dihydroxyphenylethylamine (dopamine) in the striatum of the mouse induced by drugs, Brit. J. Pharmacol. Chemotherap. 28,153. Roos, B.E., 1969, Decrease in HVA as evidence for dopaminergic receptor stimulation by apomorphine in the neostriatum of the rat, J. Pharm. Pharmacol. 21,263. Westerink, B.H.C. and J. Korf, 1976, Regional rat brain levels of 3,4-dihydroxyphenylacetic acid and homovanillic acid: concurrent fluorometric measurement and influence of drugs, European J. Pharmacol. 38, 281.
175
© E l s e v i e r / N o r t h - H o l l a n d B i o m e d i c a l Press
Short communication EFFECTS OF D R U G S ON THE F O R M A T I O N OF HOMOVANILLIC ACID IN THE RAT RETINA BEN H.C. W E S T E R I N K * a n d J A K O B K O R F **
* Laboratory for Pharmaceutical and Analytical Chemistry, Department o f Clinical Chemistry, Antonius Deusinglaan 2, and ** Department o f Biological Psychiatry, State University, Groningen, The Netherlands Received 6 A u g u s t 1976, a c c e p t e d 9 A u g u s t 1 9 7 6
B.H.C. W E S T E R I N K a n d J. K O R F , Effects o f drug on the formation o f homovanillic acid in the rat retina, E u r o p e a n J. P h a r m a c o l . 40 ( 1 9 7 6 ) 1 7 5 - - 1 7 8 . H o m o v a n i l l i c acid ( H V A ) levels were m e a s u r e d in t h e eye a n d t h e c o r p u s s t r i a t u m o f rats u n d e r n o r m a l condit i o n s a n d a f t e r d i f f e r e n t drug t r e a t m e n t s . N e u r o l e p t i c agents such as clozapine, c i s - f l u p e n t h i x o l a n d h a l o p e r i d o l i n d u c e d c o m p a r a b l e increases in H V A levels, whereas t h e n o n - n e u r o l e p t i c t r a n s - i s o m e r of f l u p e n t h i x o l was inactive in b o t h s t r u c t u r e s . A p o m o r p h i n e decreased H V A levels in t h e r e t i n a a n d t h e c o r p u s s t r i a t u m , w h i l e amp h e t a m i n e i n d u c e d a decreased H V A f o r m a t i o n in t h e r e t i n a a n d did n o t c h a n g e levels o f H V A in t h e c o r p u s s t r i a t u m . P r o b e n e c i d caused a similar p e r c e n t a g e rise of H V A in b o t h s t r u c t u r e s . M o r p h i n e a n d o x o t r e m o r i n e i n d u c e d a rise in H V A levels in the c o r p u s s t r i a t u m b u t n o t in retinal samples. H o m o v a n i l l i c acid Corpus striatum
Oxotremorine
Morphine
1. Introduction In addition to the dopamine (DA)-containing nigrostriatal and mesolimbic pathways, dopaminergic processes have been described in the retina. The retinal DA-containing neurons have been histochemically characterized and were found to be similar in morphology to amacrine cells (e.g. H~iggendal and Malmfors, 1965; Ehinger, 1966). Many centrally acting drugs e.g. neuroleptics, central stimulants, analgesics and cholinomimetics are known to influence the formation and utilization (the turnover) of DA in the central nervous system. Measurement of the levels of the two main DA metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) appeared to be a very sensitive m e t h o d to detect changes in the turnover of DA; there have thus been many studies of the formation of these metabolites in brain during various drug treatments (e.g.
Neuroleptics
Retina
Laverty and Sharman, 1965; Westerink and Korf, 1976). From studies on the formation of DOPAC and HVA in various regions of the rat brain it appeared that in general striatal and mesolimbic structures respond in a similar manner to drug treatment (Westerink and Korf, 1976). Apparently these cerebral dopaminergic systems have a comparable vulnerability to various drugs. The presence of DA in the retina however allows one to study the influence of drugs on DA metabolism in a neuronal environment which differs from the striatal and mesolimbic structures. A recently developed sensitive assay for HVA (Westerink and Korf, 1976) made it possible to investigate the formation of HVA in a single rat eye. We studied the influence of amphetamine, apomorphine, clozapine, cis-flupenthixol, trans-flupenthixol, haloperidol, morphine, oxotremorine, pargyline and probenecid on the formation of HVA in the retina. The effects of these drugs on re-
176
tinal and striatal DA-metabolism will be compared.
B.H.C. W E S T E R I N K , J. K O R F
mental values were analyzed with the Student's t-test. A p value of less than 0.05 was considered significant.
2. Materials and methods 3. Results Male albino Wistar rats of 1 7 5 - - 2 5 0 g weight (T.N.O., Zeist, The Netherlands) were used. The following drugs were used: amphetamine sulphate, apomorphine hydrochloride, clozapine, cis-flupenthixol hydrochloride, trans-flupenthixol hydrochloride, haloperidol, morphine hydrochloride, oxotremorine sesquioxalate, pargyline hydrochloride, probenecid. The doses of drugs given in Results are expressed as free base or acid. Apomorphine was dissolved in saline containing 25 mg ascorbic acid/100 ml. Haloperidol was used as a commercially prepared solution (Serenase®). Clozapine was dissolved in a drop of acetic acid and diluted with saline. All other drugs were dissolved in saline. Groups of drug-treated rats were matched with 3--4 animals injected with saline, which were considered as controls. Drugs were administered intraperitoneally and the animals were killed following the time schedules given in Results. Because HVA was found in both retinal tissue and the aqueous humor, we analyzed total eyes and considered these samples as "retinal samples". The eyes were taken out, frozen on dry ice and homogenized separately in 1.0 ml 0.4 M perchloric acid. HVA was assayed according to our previously described method (Westerink and Korf, 1976). The method is based on isolation of HVA on small Sephadex G 10 columns, followed by automated (continuous flow) fluorometric detection. Recoveries for 25 ng HVA added to cerebellar tissue were between 80 and 90%, and no corrections were made for these recoveries. Levels given for retinal as well as striatal samples are the mean of the bilateral structures, which were analyzed separately. Difficulties in obtaining correct tissue blanks prevented the determination of DOPAC (Westerink and Korf, 1976). Differences between control and experi-
Mean HVA values of saline-treated animals were (±S.E.M.) in the retinal samples: 4.3 ± 0.2 ng/eye (n = 12), and in the corpus striarum: 0.45 + 0.01 pg/g (n = 20). As after pargyline treatment (75 mg/kg, 2 h), the fluorometric assay gives values below the detection limit (1.5 ng/sample) in both striatal and retinal samples, the fluorescence recorded can be completely attributed to the presence of HVA. Table 1 shows the effects of drug treatment on HVA formation in the retina and the corpus striatum. There was significant decrease in HVA levels in both structures after apomorphine treatment (5 mg/kg, 45 min), while amphetamine (5 mg/kg, 45 min) only induced a decrease in HVA in the retinal samples. The neuroleptic agents clozapine (15 mg/kg, 2 h), cis-flupenthixol (2 mg/kg, 2 h) and haloperidol (1.0 mg/kg, 2 h) induced a significant increase of HVA levels in both retinal and striatal samples, while trans-flupenthixol (2 mg/ kg, 2 h) did not change HVA levels in either sample. After injection of morphine (20 mg/kg, 2 h) and oxotremorine (1.0 mg/kg, 60 min) significant increases of HVA levels were found only in the corpus striatum. The morphine and oxotremorine treatments were repeated in combination with probenecid (200 mg/kg, 75 min). Again no changes in HVA levels were observed in the retinal samples, although there were pronounced increases of HVA levels in striatal samples.
4. Discussion
The formation and utilization of DA in the brain appears to be very sensitive to centrally acting drugs. Many drugs, including neurolep-
H V A IN THE R E T I N A
177
TABLE 1 E f f e c t s o f various t y p e s o f drugs on H V A levels (2 S.E.M., n) in retinal samples (total eyes) and c o r p u s s t r i a t u m o f t h e rat. Drug
Dose (mg/kg)
Killed after injection (rain)
Retina (ng/eye)
Saline Amphetamine Apomorphine Clozapine Cis-flupenthixol Trans-flupenthixol Haloperidol Morphine Oxotremorine Probenecid
-5.0 5.0 15 2.0 2.0 1.0 20 1.0 200
60 45 45 120 120 120 120 120 60 120
Probenecid controls
200
75
8.0 ± 0.6 ( 7 )
1.25 -± 0.05
7)
Probenecid Morphine
200 20
75
6.7 ± 0.6 ( 5 )
2.40 ÷ 0.18
5) c
Probenecid Oxotremorine
200 1.0
75
7.7 ± 0.6 ( 6 )
1.92 ± 0.08
6) c
4.3 2.8 2.7 5.2 11.0 4.4 9.9 4.9 4.6 8.8
± 0.2 _+ 0.4 ± 0.3 ± 0.4 ± 0.7 ± 0.3 ± 0.5 ± 0.6 ± 0.3 -+ 0.6
(12) (5) (10) (9) (3) (3) (4) (8) (9) (4)
b c a c c
c
S t r i a t u m (pg/g)
0.45 0.48 0.17 1.24 2.57 0.45 2.97 1.09 0.90 1.03
± 0.01 ± 0.03 ± 0.03 ± 0.08 -± 0.18 ± 0.03 ± 0.27 ± 0.04 ± 0.09 ± 0.7
(20) (10) (5) c (7) c (3) c (3) (9) c (12) c (6) c (12)c
D i f f e r e n t f r o m c o n t r o l : Cp < 0.001; b p < 0.005; ap < 0.02.
tics (e.g. Laverty and Sharman, 1965), apomorphine (Roos, 1969), amphetamine and anorectics (Jori and Dolfini, 1974), cholino-. mimetics (Laverty and Sharman, 1965), analgesics (Sharman, 1966), LSD (Da Prada et al., 1975), antidepressants and anti-epileptics (Westerink et al., in preparation), were found to influence DA metabolism. The various mechanisms by which these drugs interact with DA turnover need further investigation. The presence of DA in the retina allows the drug-induced changes in HVA formation in the forebrain to be compared with those in a differently organized neuronal structure. We suggest that a drug acting directly on dopaminergic neurons as a result of its antagonistic or agonistic behavior at dopamine receptors, does so independently of the neuronal environment. However, an indirectly acting drug could give different responses in differently organized structures. There is considerable evidence that neuroleptics act as antagonist and apomorphine as an agonist at dopamine receptors in the brain.
Neuroleptic treatment results in an increased DA turnover in the brain and a rise in HVA levels, while apomorphine treatment causes a decreased DA turnover and decreased HVA levels (Roos, 1969). Inhibition of DA-sensitive adenylate cyclase by neuroleptics is thought to reflect their postsynaptic receptor blockade, while reversion of apomorphineinhibited synaptosomal tyrosine hydroxylase should represent their presynaptic receptor blockade (Iversen et al., 1976). These neurochemical changes induced by neuroleptics and apomorphine can be studied in striatal homogenates and are thus independent of the neuronal organization. It is therefore not surprising that neuroleptics increased HVA levels, while apomorphine reduced HVA levels in both retina and corpus striatum (table 1). The three different types of neuroleptics tested: clozapine, cis-flupenthixol and haloperidol induced comparable increases in HVA levels, whereas the non-neuroleptic transisomer of flupenthixol was inactive in both structtLres. Recently Da Prada et al. (1975) described the
178
agonistic behavior of LSD with respect to DA. The decrease of HVA levels found by these authors in both corpus striatum and retina again suggests a similar metabolic response to direct DA receptor stimulation in both structures. The decreased HVA levels in retinal samples after amphetamine treatment were not seen in corpus striatum, but the biphasic effect of this drug on DA metabolism with regard to both time and dose (e.g. Koe, 1975) could be responsible for this discrepancy. Probenecid pretreatment caused a similar percentage rise in HVA in both structures, which suggest an identical interaction with the HVA efflux in corpus striatum and retina. Morphine and oxotremorine did not induce a rise in HVA levels in the retinal samples as was seen in the corpus striatum. As combined treatment with morphine or oxotremorine with probenecid failed to elicit an additional accumulation of HVA in the retina, it seems unlikely that morphine and oxotremorine work similarly in both systems but with a different onset and duration. Morphine and oxotremorine caused identical changes of DOPAC and HVA levels in the corpus striatum (Westerink and Korf, 1976), thus we do not expect that DOPAC levels changed in retinal samples. If morphine and oxotremorine affect DA metabolism by indirect mechanisms dependent on the neuronal organisation of the corpus striatum and substantia nigra, it is conceivable that such a response is absent from retinal tissue. The results of the present study suggest that, in corpus striatum and regina, comparison of DA metabolism based on HVA levels could differentiate between drugs whose action is dependent on or independent of the connections of dopaminergic neurons with other neuronal systems.
B.H.C. WESTERINK, J. KORF
Acknowledgement The authors wish to thank Mr. Theo Mulder for expert technical assistance.
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