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The effect of valproic acid on 5-hydroxytiyptamine and 5-hydroxyindoleacetic acid concentration in hippocampal dialysates In vivo
EuropeurtJourtlalof Pharmacology,200 (1991) 11~7-169 0 1991 Elsevier Science Publishers B.V. 0014-2999/91/$03.50 ADONIS 0014299991005244
167
EJP 20865
Short communication
Peter S. Whitton and Lesiie J. Fowler Departmenrof Pharmacology,Tile School of Pharmacy,29 - 39 BrunswickSquare, London WCIN I.& UK. Received 23 April 1991, accepted 21 May 1991
The effect of valproic doleacetic acid (5-HIAA) dose-dependent increase considered with regard to
acid (100, 200 and 400 mg/kg i.p.) on extracellular 5-hydroxytryptamine (NIT) and 5-hydroxyinhas been studied using intracerebral microdialysis of the ventral hippocampus. Valproate caused a in didiysate Jc HT , but in contrast no effect on N-ii&i ievel was observed. These findings are a possible role of 5-I-IT in mediating the anticonvulsant action of valproate. Valproate; 5-HT (5-hyddroxytryptamine,
1. Xntwduction
Valproic acid (VPA) is an effective anticonvulsant
in a number of types of epilepsy. Many studies have been undertaken in an attempt to elucidate the neurochemical mode of action of this drug but as yet there is no consensus on this. An enhancement of GABA-mediated inhibition within the CNS is the most widely accepted mode of action. Previous studies have inrlicated that VPA increases the synthesis of 5=hydroxytryptamine (5-HT) and elevates brain concentration of its metabolite 5-hydroxyindoleacetic acid (5-H&4; Whitton et al., 1985). However other authors have attributed the increase in 5-HIAA or arise from a competitive inhibition by VPA on the egress of this metabolite from brain and CSF (MacMillan et al., 1987). A major drawback in all the above investigations is that measurements of either tissue or CSF concentrations of neurotransmitters and their metabolites may not reflect basal or altered concentration of neurotransmitter in the extracellular space and therefore presumed changes in postsynaptic activity. In the present study we have measured the effect of VPA on the extracellular concentrations of 5-HT and 5-HIAA using the technique of in vivo cerebral microdialysis.
Correspondence 10: P.S. Whitton, Department of Pharmacology, The School of Pharmacy, 29-34 Brunswick Square, London WClN lAX, U.K.
serotonin); Microdialysis
2. Materials and methods 2.1. Animals Male Wistar rats (280-380 g, Charles River U.K.)
were group housed in a room with a 12 h light-dark cycle and allowed food and water ad libitum. 2.2. Drug treatment VPA (Epilim, Reckitt and Colman) was dissolved in 0.9% NaCl and injected i,p. in a volume of 0.1 ml/100 g of body weight. Control animals were given 0.9% saline solution. Injections were administered between 13:00- 13:30 h. 2.3. Intracerebral
dialysis
Rats were anaesthetised with chloral hydrate BDH, 400 mg/kg i.p.1 and implanted with concentric dialysis probes into the ventral hippocampus using coordinates of Paxinos and Watson (1982; A, k3.2 mm from the intra-aural line; L, 4.75 mm from lambda; V, 8.0 mm below dura). Concentric dialysis probes were constructed as described by Whitton et al. (1990). Probes were implanted directly without the use of a guide assembly and perfusion was started 18 h later using artificial CSF of a composition previously described (Whitton et al., 1990) containing 1 PM of the 5-I-IT reuptake inhibitor citalopram, at 0.5 gl/min (Harvard microiitre perfusion pumps). In vitro recoveries of 5-HT and 5-HIAA across the dialysis probe membrane were 26-Z% and 19-21% respectively and dialysate con-
~e~~~~~~~o~s of both stlbstences were corrected for these te samples were collected every 30 min min period of stabilisation. Five consecre collected prior to VPA or saline and 5-HIAA were determined in igh pressure liquid chromatography as 9393 except that the momM acetate; 35 mM citrate anol. 0.06 mM sodium octyl
to one-way analysis of variance ificansi: of drug effect was assessed using the multiple ranges test.
ata was subjected
Figure la shows the effect of VPA on dialysate concentration of 5-HT. VPA caused a dose-dependent increase in extracellular 5-HT concentration. On the a
Etfect of Walproete on 5-HT ?iQ
“PA
400
mQlhQ
v VPA
200
mglko
* VPA
100
mglkg
.
1
1
SALINE
z L
z
360
0
210
I
3
6
12
0
Sample No.
Effect of
b
Walproate on 5-HIAA
720
400
mp/kg
VPA
200
m#lkg
* VPA
100
mplkg
#a SALINE
160 I 610
0
a VPA -
3
6
9
VPA had no discernable effect on extracellular 5-HIAA concentration (fig. lb), a rather surprising observation in the light of previous studies (see Introduction).
other hand
Basal concentrations of 5-HT and 5-HIAA in hippocampal dialysates were 0.17 & 0.04 and 12.95 k 4.03 pmol/lO ~1 respectivley (mean of three successive pietreatment samples from each rat t S.D., n = 20 rats). The present data demonstrate that VPA increases extracellular 5-HT concentration (fig. la) and may therefore be expected to enhance serotonergic transmission. These data do not distinguish between the possibilities that VPA increases 5-HT turnover or has a direct action on 5-HT release. An effect of VPA on 5-HT turnover would however be consistent with prhious observations (Whitton et al., 1985). The lack of effect of VPA on extracellular 5-HIAA concentration was unexpected in the light of previous observations (Whitton et al., 1985; MacMillan et al., 1987). This may reflect the differerlce in methodologies employed in this and the above cited studies, or could be due to a regionally specific effect of VPA on 5-H!AA concentration as was observed by Shukla (1985). It is, however, quite possible that this is due to the presence of the 5-HT reuptake inhibitor citalopram in the perfusate thereby preventing reuptake aad subsequent deamination of 5-HT intracellularly. The relevance of enhanced serotonergic function with respect to VPA and its antiepileptic efficacy is unclear. It can be seen from fig. la that the duration over which VPA elevated dialysate 5-HT was quite short. However, a number of studies using comparable doses of VPA have observed relatively short periods of anticonvulsant activity of up to 2 h or less (e.g. Frey and Loscher, 1976). Against pentylenetetrazol-induced seizures Lazarova et al. (1983) considered that 5-HT may play a role in mediating the anticonvulsant effect of VPA but using the same seizure model Abed (1990) did not find any such role. However, increased serotonergic function has been implicated in the therapy of certain epileptic conditions such as myoclonus (Chadwick et al., 19751, a condition in which VPA is effective. The possible role of 5-HT in mediating the anticonvulsant action of VPA may therefore require the use of an appropriate model of epilepsy.
12
Sample No. Fig. 1. Effect of valproate (100. 200 and 400 mg/kg i.p.) on hippocampal dialysate .5-HT concentration (a) and on S-HIAA concentration (b). Data are the meanskS.E. of 4-6 rats in each group. * Denotes pairs of groups significantly different from controls at 0.05 level.
References Abed, W.T.. 1990. Anticonvulsant activity of di-n-propylacetate and brain monoamine metabolism in the rat, Clin. Exp. Pharmacol. Physiol. 17, 11.
169 Chadwick, D., R. Harris, P. Jenner, E.H. Reynolds and C.D. Marsden, 1975, Manipulation of brain serotonin in the treatment of myoclonus, Lance1 2, 434. Frey, H.-H. and W. Loscher, 1976. Di-n-propylacetic acid-profile of anticonvulsant activity in mice, Arzneim. Forsch./Drug Res. 26, 299. Hutson, P.H., G.S. Sarna, M.T. O’Connell and G. Curzon. 1989, Hippocampal 5-HT synthesis and release in vivo is decreased by infusion of I-OHDPAT into the nucleus raphe dorsalis. Neurosci. Lett. 100, 276. Lazarova, M., C. Benedotti and R. Samanin, 1983, Studies on the role of serotonin in different regions of the rat central nervous system on pentylenetetrazol-induced seizures and the effect of di-n-propylacetate, Naunyn-Schmiedeb. Arch. Pharmacol. 338, 655. MacMillan, V.. J. Leake, T. Chung and M. Bovell, 1987, The effect
of valproic acid on the %hydroyindoleactic, homovanillic and tactic acid levels of cerebrospinal fluid, Brain Res. 420, 268. Paxinos, C. and C. Watson, 1982, The Rat Brain (Academic Press, New York). Shukla. G.S., 19*;5, Combined lithium and valproate treatment and subsequent withdrawal: serotonergic mechanism of their interaction in discrete brain regions, Prog. Nemo-Psychopharmacol. Biol. Psychiat. 9, 153. Whitton, P.S., D. Oreskovic, B. Jernej and M. Bulat, 1985, Effect of valproic acid on 5-hydroxytryptamine turnover in mouse brain, J. Pharm. Pharmacol. 37, 199. Whitton. P.S., G.S. Sarna, M.T. O’Connell and G. Curzon, 1990, The effect of the novel antidepressant tianep:ine on S-hydroxytryptamine concentration in rat hippocampal dialysates in vivo. Neuropharmacology 30. 1.
167
EJP 20865
Short communication
Peter S. Whitton and Lesiie J. Fowler Departmenrof Pharmacology,Tile School of Pharmacy,29 - 39 BrunswickSquare, London WCIN I.& UK. Received 23 April 1991, accepted 21 May 1991
The effect of valproic doleacetic acid (5-HIAA) dose-dependent increase considered with regard to
acid (100, 200 and 400 mg/kg i.p.) on extracellular 5-hydroxytryptamine (NIT) and 5-hydroxyinhas been studied using intracerebral microdialysis of the ventral hippocampus. Valproate caused a in didiysate Jc HT , but in contrast no effect on N-ii&i ievel was observed. These findings are a possible role of 5-I-IT in mediating the anticonvulsant action of valproate. Valproate; 5-HT (5-hyddroxytryptamine,
1. Xntwduction
Valproic acid (VPA) is an effective anticonvulsant
in a number of types of epilepsy. Many studies have been undertaken in an attempt to elucidate the neurochemical mode of action of this drug but as yet there is no consensus on this. An enhancement of GABA-mediated inhibition within the CNS is the most widely accepted mode of action. Previous studies have inrlicated that VPA increases the synthesis of 5=hydroxytryptamine (5-HT) and elevates brain concentration of its metabolite 5-hydroxyindoleacetic acid (5-H&4; Whitton et al., 1985). However other authors have attributed the increase in 5-HIAA or arise from a competitive inhibition by VPA on the egress of this metabolite from brain and CSF (MacMillan et al., 1987). A major drawback in all the above investigations is that measurements of either tissue or CSF concentrations of neurotransmitters and their metabolites may not reflect basal or altered concentration of neurotransmitter in the extracellular space and therefore presumed changes in postsynaptic activity. In the present study we have measured the effect of VPA on the extracellular concentrations of 5-HT and 5-HIAA using the technique of in vivo cerebral microdialysis.
Correspondence 10: P.S. Whitton, Department of Pharmacology, The School of Pharmacy, 29-34 Brunswick Square, London WClN lAX, U.K.
serotonin); Microdialysis
2. Materials and methods 2.1. Animals Male Wistar rats (280-380 g, Charles River U.K.)
were group housed in a room with a 12 h light-dark cycle and allowed food and water ad libitum. 2.2. Drug treatment VPA (Epilim, Reckitt and Colman) was dissolved in 0.9% NaCl and injected i,p. in a volume of 0.1 ml/100 g of body weight. Control animals were given 0.9% saline solution. Injections were administered between 13:00- 13:30 h. 2.3. Intracerebral
dialysis
Rats were anaesthetised with chloral hydrate BDH, 400 mg/kg i.p.1 and implanted with concentric dialysis probes into the ventral hippocampus using coordinates of Paxinos and Watson (1982; A, k3.2 mm from the intra-aural line; L, 4.75 mm from lambda; V, 8.0 mm below dura). Concentric dialysis probes were constructed as described by Whitton et al. (1990). Probes were implanted directly without the use of a guide assembly and perfusion was started 18 h later using artificial CSF of a composition previously described (Whitton et al., 1990) containing 1 PM of the 5-I-IT reuptake inhibitor citalopram, at 0.5 gl/min (Harvard microiitre perfusion pumps). In vitro recoveries of 5-HT and 5-HIAA across the dialysis probe membrane were 26-Z% and 19-21% respectively and dialysate con-
~e~~~~~~~o~s of both stlbstences were corrected for these te samples were collected every 30 min min period of stabilisation. Five consecre collected prior to VPA or saline and 5-HIAA were determined in igh pressure liquid chromatography as 9393 except that the momM acetate; 35 mM citrate anol. 0.06 mM sodium octyl
to one-way analysis of variance ificansi: of drug effect was assessed using the multiple ranges test.
ata was subjected
Figure la shows the effect of VPA on dialysate concentration of 5-HT. VPA caused a dose-dependent increase in extracellular 5-HT concentration. On the a
Etfect of Walproete on 5-HT ?iQ
“PA
400
mQlhQ
v VPA
200
mglko
* VPA
100
mglkg
.
1
1
SALINE
z L
z
360
0
210
I
3
6
12
0
Sample No.
Effect of
b
Walproate on 5-HIAA
720
400
mp/kg
VPA
200
m#lkg
* VPA
100
mplkg
#a SALINE
160 I 610
0
a VPA -
3
6
9
VPA had no discernable effect on extracellular 5-HIAA concentration (fig. lb), a rather surprising observation in the light of previous studies (see Introduction).
other hand
Basal concentrations of 5-HT and 5-HIAA in hippocampal dialysates were 0.17 & 0.04 and 12.95 k 4.03 pmol/lO ~1 respectivley (mean of three successive pietreatment samples from each rat t S.D., n = 20 rats). The present data demonstrate that VPA increases extracellular 5-HT concentration (fig. la) and may therefore be expected to enhance serotonergic transmission. These data do not distinguish between the possibilities that VPA increases 5-HT turnover or has a direct action on 5-HT release. An effect of VPA on 5-HT turnover would however be consistent with prhious observations (Whitton et al., 1985). The lack of effect of VPA on extracellular 5-HIAA concentration was unexpected in the light of previous observations (Whitton et al., 1985; MacMillan et al., 1987). This may reflect the differerlce in methodologies employed in this and the above cited studies, or could be due to a regionally specific effect of VPA on 5-H!AA concentration as was observed by Shukla (1985). It is, however, quite possible that this is due to the presence of the 5-HT reuptake inhibitor citalopram in the perfusate thereby preventing reuptake aad subsequent deamination of 5-HT intracellularly. The relevance of enhanced serotonergic function with respect to VPA and its antiepileptic efficacy is unclear. It can be seen from fig. la that the duration over which VPA elevated dialysate 5-HT was quite short. However, a number of studies using comparable doses of VPA have observed relatively short periods of anticonvulsant activity of up to 2 h or less (e.g. Frey and Loscher, 1976). Against pentylenetetrazol-induced seizures Lazarova et al. (1983) considered that 5-HT may play a role in mediating the anticonvulsant effect of VPA but using the same seizure model Abed (1990) did not find any such role. However, increased serotonergic function has been implicated in the therapy of certain epileptic conditions such as myoclonus (Chadwick et al., 19751, a condition in which VPA is effective. The possible role of 5-HT in mediating the anticonvulsant action of VPA may therefore require the use of an appropriate model of epilepsy.
12
Sample No. Fig. 1. Effect of valproate (100. 200 and 400 mg/kg i.p.) on hippocampal dialysate .5-HT concentration (a) and on S-HIAA concentration (b). Data are the meanskS.E. of 4-6 rats in each group. * Denotes pairs of groups significantly different from controls at 0.05 level.
References Abed, W.T.. 1990. Anticonvulsant activity of di-n-propylacetate and brain monoamine metabolism in the rat, Clin. Exp. Pharmacol. Physiol. 17, 11.
169 Chadwick, D., R. Harris, P. Jenner, E.H. Reynolds and C.D. Marsden, 1975, Manipulation of brain serotonin in the treatment of myoclonus, Lance1 2, 434. Frey, H.-H. and W. Loscher, 1976. Di-n-propylacetic acid-profile of anticonvulsant activity in mice, Arzneim. Forsch./Drug Res. 26, 299. Hutson, P.H., G.S. Sarna, M.T. O’Connell and G. Curzon. 1989, Hippocampal 5-HT synthesis and release in vivo is decreased by infusion of I-OHDPAT into the nucleus raphe dorsalis. Neurosci. Lett. 100, 276. Lazarova, M., C. Benedotti and R. Samanin, 1983, Studies on the role of serotonin in different regions of the rat central nervous system on pentylenetetrazol-induced seizures and the effect of di-n-propylacetate, Naunyn-Schmiedeb. Arch. Pharmacol. 338, 655. MacMillan, V.. J. Leake, T. Chung and M. Bovell, 1987, The effect
of valproic acid on the %hydroyindoleactic, homovanillic and tactic acid levels of cerebrospinal fluid, Brain Res. 420, 268. Paxinos, C. and C. Watson, 1982, The Rat Brain (Academic Press, New York). Shukla. G.S., 19*;5, Combined lithium and valproate treatment and subsequent withdrawal: serotonergic mechanism of their interaction in discrete brain regions, Prog. Nemo-Psychopharmacol. Biol. Psychiat. 9, 153. Whitton, P.S., D. Oreskovic, B. Jernej and M. Bulat, 1985, Effect of valproic acid on 5-hydroxytryptamine turnover in mouse brain, J. Pharm. Pharmacol. 37, 199. Whitton. P.S., G.S. Sarna, M.T. O’Connell and G. Curzon, 1990, The effect of the novel antidepressant tianep:ine on S-hydroxytryptamine concentration in rat hippocampal dialysates in vivo. Neuropharmacology 30. 1.