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12 in rats
Brain Research, 594 (1992) 323-326 © 1992 Elsevier Science Publishers B.V. All rights reserved 0006-8993/92/$05.00
323
BRES 25378
Dextromethorphan antagonizes the acute depletion of brain serotonin by p-chloroamphetamine and H75/12 in rats M a r k G. H e n d e r s o n and R a y W~ Fuller Lilly Research Laboratories, Eli Lilly and Company, Indlanapohs, IN 46285 (USA) (Accepted 30 June 1992)
Key words: Parachloroamphetamine; Dextromethorphan; H75/12; Neurotoxicity; Serotonin
A role for calcium in p-chloroamphetamine-induced neurotoxicity has been inferred previously from protective effects of dextromethorphan. We found that dextromethorphan reduces rat brain concentrations of 5-hydroxs,indoleacetic acid and blocks the acute, non-neurotoxic depletion of brain serotonin by p-chloroamphetamine and by H75/12, Inhibition of the membrane transporter on brain serotonin neurons by dextromethorphan in vivo might explain its protective effect against p-chloroamphetamine neurotoxicity.
p-Chloroamphetamine (PCA) at single doses of approximately 10 mg/kg and higher causes long-lasting depletion of brain serotonin in rats 24'2s. Accompanying the decrease in brain serotonin content are persistent decreases in other ~arameters specifically associated with brain serotonin neurons. For example, there are decreases in (a) the concentration of 5-hydroxyindoloacetic acid (SHIAA) and in the accumulation of 5HIAA after probenecid administration to block its effiux from brain, (b) tryptophan hydroxylase activity measured in vitro and in vivo, (c) the number of serotonin-containing nerve fibers identified histochemically, (d) serotonin uptake capacity, (e) binding sites for radioligands that label the serotonin uptake carrier, and (f) the number of neurons projecting from the dorsal raphe that are retrogradely labeled with fluorogold injected into terminal regions 15a6aT'2°'24'~5. These findings constitute strong evidence that high doses of PCA are neurotoxic toward some serotonm neurons in rat brain, and histologic evidence for degenerating axons at short times after PCA injection has been reported 6. Recently, Finnegan et al. 7 reported that dextromethorphan protects against the neurotoxic effects of PCA in rats. They measured serotonin concentration in brain regions 10 days after a single dose of PCA
and observed that dextromethorphan pretreatment produced a dose-related antagonism of the serotonin depletion. Finnegan et al. 7 interpreted their findings in light of dextromethorphan's reported ability to block glutamate-induced neurotoxicity in neuronal cell culture 4, effects possibly related to the ability of dextromethorphan to block calcium channels in vitro "~. Another, more likely, explanation for dextromethorphan antagonism of PCA effects was not considered. Dextromethorphan, like some other opioid drugs, is a relatively potent inhibitor of the serotonin uptake carrier in vitro ~'tg'2r'. There is evidence that dextromethorphan inhibits the uptake carrier on brain serotonin neurons in vivo. t'26. The depletion of brain serotonin by PCA is dependent upon the serotonin uptake carrier and is blocked by inhibitors of that uptake carrier ~'ms'2s. Indeed, blockade of PCA-induced depletion of brain serotonin has been a standard method for assessing the in vivo efficacy of inhibitors of the serotonin uptake carrier 12.2m'2s. To test the possibility that inhibition of the uptake carrier on serotonin neurons accounts for the antagonism of PCA neurotoxicity reported by Finnegan et al. 7, we have determined the ability of dextromethorphan to antagonize the short-term, non.neurotoxic depletion of brain serotonin by PCA and by H75/12
Correspondence: R.W. Fuller, CNS Research, Lilly Research Laboratories, Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285, USA.
324 (4-methyl-a-ethyl-m-tyramine), The latter compound is another amphetamine analog like PCA but causes only transient depletion of brain serotonin without being neurotoxic TM. The acute depletion of brain serotonin by PCA in rats is fully reversible by drug treatment :3, i.e,, the initial serotonin depletion at 2 h does not represent a neurotoxic effect of PCA. Inhibitors of the serotonin uptake carrier block both the acute, reversible and the long-term, neurotoxic depletion of serotonin by PCAs. Male Sprague-Dawley rats (190-210 g) from Charles River Breeding Laboratories, Portage, MI) were housed in a temperature- and light-controlled room with food and water freely available. (±)p-Chloroamphetamine hydrochloride (Regis Chemical Company, Morton Grove, IL) was injected at a dose of 10 mg/kg i.p. 2 h before rats were killed. H75/12, 4methyl-a-ethyl-m-tyramine (Labkemi AB, Stockholm, Sweden) was injected at a dose of 40 mg/kg i.p. 2 h before rats were killed. Dextromethorphan hydrobromide (Sigma Chemical Company, St. Louis, Me) was injected at doses of 25 or 75 mg/kg i.p. 140 minutes before rats were killed. Whole brains were removed after rats were decapitated and were then frozen on dry-ice and stored at -70°C prior to analysis. Serotonin and 5HIAA concentrations were determined by liquid chromatography with electrochemical detection ~c~. Statistical analyses wore done by analysis of variance followed by Tukey's post hoc test, Fig. I shows that PCA caused 47% depletion of serotonin concentration in whole brain 2 h after its injection into rats. This depletion was totally prevented by pretreatment with the 75 mg/kg dose of dextromethorphan, although the 25 mg/kg dose of de×tromethorphan was ineffective. H75/12 caused 56% depletion of brain serotonin at 2 h, and this depletion was antagonized significantly by both doses of dextromethorphan. Table I shows that dextromethorphan decreased whole brain concentrations of the serotonin metabo. lite, 5HIAA. The decrease was statistically significant at both doses of dextromethorphan. At the same time, serotonin concentration was not decreased (see Fig. 1, left panel). Our findings suggest that dextromethorphan at doses in the range of 25-75 mg/kg i.D. in rats inhibits the uptake carrier on brain serotonin neurons, agreeing with earlier in rive data published by Ahtee:. Our results show that dextromethorphan antagonized the short-term, non-neurotoxic serotonin depletion by PCA, just as Finnegan et al.7 had reported it to antagonize the long-term, neurotoxic effect. In addition, our results show that dextromethorphan antagonized H75/
CONTROL
H75/12
PCA i
t
E o O
E
I=
0 25 76
0
26 T8
O 28 78
DOSE OF DEXTROMETHORPHAN(nlWttg, i.p.) "slgnlflunUy different from control p,(O.06 § slgnlfl©llntlydifferent from H7[I/12or POA alone p
Fig. I. Dextromethorphan pretreatment antagonizes brain serotonin depletion by PCA and by H75/12 in rats. PCA hydrochloride (10 mg/kg i.p.) and H75/12 (40 mg/kg i.p,) were injected 20 rain after dextromethorphan (25 or 75 mg/k8 i.p.) and 2 h before rats were killed. Mean values + standard errors for 6 rats per group are shown. Asterisks indicate significant depletion of brain serotonin compared to the corresponding control group, and (§) indicate significant protection by dextromethorphan (P < 0.05).
12, another serotonin-depleting agent which is not neurotoxic but causes transient depletion of brain serotonin that is carrier-dependent as are the effects of PCA. The decrease in 5tllAA/serotonin ratio caused by de×tromethorphan (Table 1) replicates the findings of Ahtee t and further indicates that dextromethorphan inhibits the serotonin uptake carrier at these doses, as a decrease in 5HIAA/serotonin ratio is well known to occur with serotonin uptake inhibitorst4. The mechanism for this decrease is thought to be a reduction in serotonin synthesis and turnover secondary to increased concentrations of serotonin in the synaptic cleft when the membrane uptake carrier is blocked. TABLE I
D~'tromethorphan lowers brain 5HI/IA concentration m rats Dextromethorphan was injected 140 min before rats were killed. Mean values+standard errors for 6 rats per group are shown. Asterisk indicate significant drug effect (P < 0.05).
Dose of dextrom,,thorphan (mg / kg, i.p)
5HIAA in whole brain (nmoles /g)
0 25 75
2.17±0.12 1 69±0.07 * (-22%) 1.64±0.15 * (-25%)
325
Various measures, including 5-hydroxytryptophan accumulation after decarboxylaxe inhibition and incorporation of radioactive tryptophan into 5-hydroxyindoles, show a reduction in serotonin synthesis when uptake is inhibited t4. As an electrophysiological correlate of these neurochemical measures, the firing of serotonin neurons in raphe areas is reduced in the presence of uptake inhibitors 5'23. Dextromethorphan is able to inhibit the serotonin uptake carrier in vivo because it has relatively high affinity for that uptake carrier. The IC5o concentration for dextromethorphan as an inhibitor of serotonin uptake by blood platelets in vitro is 120 nM t. In contrast, dextromethorphan inhibits calcium uptake in PC12 cells in vitro with an IC5o of I01 /zM 3, a concentration nearly 3 orders of magnitude higher. These considerations and our current findings suggest the most likely explanation for the dextromethorphan antagonism of serotonin neurotoxicity 10 days after PCA injection, as described by Finnegan et al. 7, is inhibition of the serotonin uptake carrier by dextromethorphan. Dextromethorphan has been reported to have neuroprotective effects in models of hypoxia-ischemia in laboratory animals in vivo 2z2~. There is no reason to suspect that inhibition of serotonin uptake is involved in those actions of dextromethorphan. Whether the effects of dextromethorphan in these models are due to antagonism of excitatory amino acid receptors, inhibition of calcium entry, actions on opioid receptors, or some other mechanism, the possibility must be considered that the same mechanisms account for its protection against PCA neurotoxicity, But because it is already well known that serotonin uptake inhibitors block the neurotoxic actions of PCA~'ts'2s, the protective effects of dextromethorphan do not necessarily implicate excitatory amino acid receptors or calcium entry in the neurotoxic actions of PCA. If agents that block calcium entry selectively were found to protect against PCA neurotoxicity, that could implicate altered calcium homeostasis in the neurotoxic actions of PCA. I Ahtee, L., Dextromethorphan inhibits 5.hydroxytryptamine uptake by human blood platelets and decreases 5-hydroxyindoleacetic acid content in rat brain, J. Pharm. Pharmacol., 27 (1975) 177-180. 2 Carlsson, A., Corrodi, H., Fuxe, K. and H0kfeit, T., Effect of antidepressant drugs on the depletion of intraneuronal brain 5-hydroxytryptamine stores caused by 4-methyl.a-ethyl-metatryamine, Ear. J. PharmacoL, 5 (1969) 357-366. 3 Carpenter, C.L., Marks, S.S., Watson, D.L. and Greenberg, D.A., Dextromethorphan and dextrorphan as calcium channel antagonists, Brain Res., 439 (1988) 372-375. 4 Choi, D.W., Dextrorphan and dextromethorphan attenuate glutamate neurotoxicity, Brain Res., 403 (1987) 333-336. 5 Clemens, J.A., Sawyer, B.D. and Cerimele, B., Further evidence
that serotonin i,~ a neurotransmitter involved in the control of prolactin secretion, Endocrinology, 100 (1977) 692-698. 6 Commins, D.L., Axt, K.J., Vosmer, G. and Seiden, L.S., Endogenously produced 5,6-dihydroxytryptamme may medmte the neurotoxic effects el para-chloroamphetamine, Brain Res., 419 (1987) 253-261. 7 Finnegan, K.T, Kerr, J.T. and Langston, J.W., Dextromethorphan protects against the neurotox~c effects of p-chloroamphetamine in rats, Brain Res., 558 (1991) 109-1il. 8 Fuller, R.W., Effects of p-chioroamphetamine on brain serotonin neurons, Neurochem. Res., 17 (1992)449-456. 9 Fuller, R.W., Mechanism by which uptake inhibitors antagomze p-chloroamphetamine-induced depletion of brain serotonin, Neurochem. Res., 5 (1980) 241-245. 10 Fuller, R.W. and Perry, K.W., Effects of buspirone and its metabolite, 1-(2-pyrimidinyl)piperazine,on brain monoammes and their metabolites w rats, J. Pharmacol. Exp. Ther., 248 (1989) 50-56. 11 Fuller, R.W., Perry, K.W. and Baker, J.C., Duration of the effects of a-ethyl-4-methyi-m-tyramine, (H75/12)on bram 5-hydroxyindole concentratiuns in rats, J. Pharm. PharmacoL, 28 (1976) 649-650. 12 Fuller, R.W., Perry, K.W. and Moiloy, B.B., Effect of 3-(p-trifluoro-methylphenoxy)-N-methyl-3-phenylpropylamine on the depletion of brain serotonin by 4-chloroamphetamine, J. Pharmacol. Exp. Ther,, 193 0975)796-803. 13 Fuller, R.W., Perry, K.W. and Molloy, B.B., Reversible and irreversible phases of serotonin depletion by 4-chloroamphetamine, Eur. J. PharmacoL, 33 (1975) 119-124. 14 Fuller, R.W. and Wong, D.T., Serotonin reuptake blockers in vitro and in rive, J. Chn. Psychopharmacol., 7 (1987) 36S-43S. 15 Harvey, J.A., McMaster, S.E. and Fuller, R.W., Comparison between the neurotoxic and serotonin-depleting effects of various halogenated derivatives of amphetamine in the rat, J. PharmacoL Exp. Ther., 202 (1977) 581-589. 16 Hashimoto, K. and Goromaru, T., High-affinity [~H]6-nitroquipazine binding sites in rat brain, Ear, J, Phamtacol., 180 (1990) 273-281. 17 Mamounas, L.A, and Molliver, M.E., Evidence for dual serotonergic projections to neocortex: axons from the dorsal and median raphe nuclei are differentially vulnerabl~ to the neurotoxin pchloroamphetamine (PCA), Exp. NearoL, 102 (1988) 23-36. 18 Meek, J.L., Fuxe, K. and Carlsson, A., Blockade of p.chloromethamphetamine induced 5-hydro~trypt:~mine depletion by chlorimipramine, chlorpheniramine and meperidine, Biochem. Pharmacol., 20 (I 971) 707-709. 19 Moffat, J,A. and Jhamandas, K., Effects of acute and chronic methadone treatment of the uptake of 31ol-5-hydroxytryplamine in rat hypoth:~lamu~:dices, Ear. J. Pharmacol., 36 (1976) 289-297. 20 Molliver, M.E., Berger, U.V., Mamounas, L.A., MoUiver, D.C., O'Hearn, E. and Wilson, M.A., Neurotoxicity of MDMA and related compounds: anatomic studies, Ann. N.Y. Acad. Sc'i., 600 (1990) 640-664. 21 Perrone, M.H., Luttinger, D., Hamel, L.T., Fritz, P.M., Ferraini, g. and Haubrich, D.R., In vivo assessment of napamezole, an alpha-2 adrenoceptor antagonist and monoamine re-uptake inhibitors, J. Pharmacol. Exp. Ther., 254 (1990) 476-483. 22 Prince, D.A. and Feeser, H.R., Dextromethorphan protects against cerebral infarction in a rat model of hypoxia-isehemia, Neurosci. Lett., 85 (1988) 291-296. 23 Rigdon, G.C. and Wang, C.M., Serotonin uptake blockers inhibit the firing of presumed serotonergic dorsal raphe neurons in vitro, Drag DeL'. Res., 22 (199t) 135-140. 24 Sanders-Bush, E., Bushing, J.A. and Sulser, F., Long-term effects of p-chloroamphetamine on tryptophan hydroxylase activity and on the !evels of 5.hydroxytryptamine and 5.hydro~indoleacetic acid in brain, Fur. J. Pharmacol., 20 (1972) 385-388. 25 Sanders-Bush, E. and Steranka, L.R., Immediate and long-term effects of p.chloroamphetamine on brain amines, Am~. N.Y. Acad. Sct., 305 (1978) 208-221.
326 26 Sinclair, J.G. and Lo, G.F., The blockade of serotonin uptake into synaptosomes: relationship to an interaction with monoamine oxidase inhibitors, Can. I. PhysioL Pharmacol., 55 (1977) 180-187. 27 Steinberg, G.K., Saleh, J. and Kunis, D., Delayed treatment with dextromethorphan and dextrorphan reduces cerebral damage after transient focal inschemia, Neurosct. Left, 89 (1988) 193-197.
28 Thomas, D.R., Nelson, D.R. and Johnson, A.M., Biochemical effects of the antidepressant paroxetine, a specific 5-hydroxytryptamine uptake inhibitor, Psychopharmacology, 93 (1987) 193200.
323
BRES 25378
Dextromethorphan antagonizes the acute depletion of brain serotonin by p-chloroamphetamine and H75/12 in rats M a r k G. H e n d e r s o n and R a y W~ Fuller Lilly Research Laboratories, Eli Lilly and Company, Indlanapohs, IN 46285 (USA) (Accepted 30 June 1992)
Key words: Parachloroamphetamine; Dextromethorphan; H75/12; Neurotoxicity; Serotonin
A role for calcium in p-chloroamphetamine-induced neurotoxicity has been inferred previously from protective effects of dextromethorphan. We found that dextromethorphan reduces rat brain concentrations of 5-hydroxs,indoleacetic acid and blocks the acute, non-neurotoxic depletion of brain serotonin by p-chloroamphetamine and by H75/12, Inhibition of the membrane transporter on brain serotonin neurons by dextromethorphan in vivo might explain its protective effect against p-chloroamphetamine neurotoxicity.
p-Chloroamphetamine (PCA) at single doses of approximately 10 mg/kg and higher causes long-lasting depletion of brain serotonin in rats 24'2s. Accompanying the decrease in brain serotonin content are persistent decreases in other ~arameters specifically associated with brain serotonin neurons. For example, there are decreases in (a) the concentration of 5-hydroxyindoloacetic acid (SHIAA) and in the accumulation of 5HIAA after probenecid administration to block its effiux from brain, (b) tryptophan hydroxylase activity measured in vitro and in vivo, (c) the number of serotonin-containing nerve fibers identified histochemically, (d) serotonin uptake capacity, (e) binding sites for radioligands that label the serotonin uptake carrier, and (f) the number of neurons projecting from the dorsal raphe that are retrogradely labeled with fluorogold injected into terminal regions 15a6aT'2°'24'~5. These findings constitute strong evidence that high doses of PCA are neurotoxic toward some serotonm neurons in rat brain, and histologic evidence for degenerating axons at short times after PCA injection has been reported 6. Recently, Finnegan et al. 7 reported that dextromethorphan protects against the neurotoxic effects of PCA in rats. They measured serotonin concentration in brain regions 10 days after a single dose of PCA
and observed that dextromethorphan pretreatment produced a dose-related antagonism of the serotonin depletion. Finnegan et al. 7 interpreted their findings in light of dextromethorphan's reported ability to block glutamate-induced neurotoxicity in neuronal cell culture 4, effects possibly related to the ability of dextromethorphan to block calcium channels in vitro "~. Another, more likely, explanation for dextromethorphan antagonism of PCA effects was not considered. Dextromethorphan, like some other opioid drugs, is a relatively potent inhibitor of the serotonin uptake carrier in vitro ~'tg'2r'. There is evidence that dextromethorphan inhibits the uptake carrier on brain serotonin neurons in vivo. t'26. The depletion of brain serotonin by PCA is dependent upon the serotonin uptake carrier and is blocked by inhibitors of that uptake carrier ~'ms'2s. Indeed, blockade of PCA-induced depletion of brain serotonin has been a standard method for assessing the in vivo efficacy of inhibitors of the serotonin uptake carrier 12.2m'2s. To test the possibility that inhibition of the uptake carrier on serotonin neurons accounts for the antagonism of PCA neurotoxicity reported by Finnegan et al. 7, we have determined the ability of dextromethorphan to antagonize the short-term, non.neurotoxic depletion of brain serotonin by PCA and by H75/12
Correspondence: R.W. Fuller, CNS Research, Lilly Research Laboratories, Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285, USA.
324 (4-methyl-a-ethyl-m-tyramine), The latter compound is another amphetamine analog like PCA but causes only transient depletion of brain serotonin without being neurotoxic TM. The acute depletion of brain serotonin by PCA in rats is fully reversible by drug treatment :3, i.e,, the initial serotonin depletion at 2 h does not represent a neurotoxic effect of PCA. Inhibitors of the serotonin uptake carrier block both the acute, reversible and the long-term, neurotoxic depletion of serotonin by PCAs. Male Sprague-Dawley rats (190-210 g) from Charles River Breeding Laboratories, Portage, MI) were housed in a temperature- and light-controlled room with food and water freely available. (±)p-Chloroamphetamine hydrochloride (Regis Chemical Company, Morton Grove, IL) was injected at a dose of 10 mg/kg i.p. 2 h before rats were killed. H75/12, 4methyl-a-ethyl-m-tyramine (Labkemi AB, Stockholm, Sweden) was injected at a dose of 40 mg/kg i.p. 2 h before rats were killed. Dextromethorphan hydrobromide (Sigma Chemical Company, St. Louis, Me) was injected at doses of 25 or 75 mg/kg i.p. 140 minutes before rats were killed. Whole brains were removed after rats were decapitated and were then frozen on dry-ice and stored at -70°C prior to analysis. Serotonin and 5HIAA concentrations were determined by liquid chromatography with electrochemical detection ~c~. Statistical analyses wore done by analysis of variance followed by Tukey's post hoc test, Fig. I shows that PCA caused 47% depletion of serotonin concentration in whole brain 2 h after its injection into rats. This depletion was totally prevented by pretreatment with the 75 mg/kg dose of dextromethorphan, although the 25 mg/kg dose of de×tromethorphan was ineffective. H75/12 caused 56% depletion of brain serotonin at 2 h, and this depletion was antagonized significantly by both doses of dextromethorphan. Table I shows that dextromethorphan decreased whole brain concentrations of the serotonin metabo. lite, 5HIAA. The decrease was statistically significant at both doses of dextromethorphan. At the same time, serotonin concentration was not decreased (see Fig. 1, left panel). Our findings suggest that dextromethorphan at doses in the range of 25-75 mg/kg i.D. in rats inhibits the uptake carrier on brain serotonin neurons, agreeing with earlier in rive data published by Ahtee:. Our results show that dextromethorphan antagonized the short-term, non-neurotoxic serotonin depletion by PCA, just as Finnegan et al.7 had reported it to antagonize the long-term, neurotoxic effect. In addition, our results show that dextromethorphan antagonized H75/
CONTROL
H75/12
PCA i
t
E o O
E
I=
0 25 76
0
26 T8
O 28 78
DOSE OF DEXTROMETHORPHAN(nlWttg, i.p.) "slgnlflunUy different from control p,(O.06 § slgnlfl©llntlydifferent from H7[I/12or POA alone p
Fig. I. Dextromethorphan pretreatment antagonizes brain serotonin depletion by PCA and by H75/12 in rats. PCA hydrochloride (10 mg/kg i.p.) and H75/12 (40 mg/kg i.p,) were injected 20 rain after dextromethorphan (25 or 75 mg/k8 i.p.) and 2 h before rats were killed. Mean values + standard errors for 6 rats per group are shown. Asterisks indicate significant depletion of brain serotonin compared to the corresponding control group, and (§) indicate significant protection by dextromethorphan (P < 0.05).
12, another serotonin-depleting agent which is not neurotoxic but causes transient depletion of brain serotonin that is carrier-dependent as are the effects of PCA. The decrease in 5tllAA/serotonin ratio caused by de×tromethorphan (Table 1) replicates the findings of Ahtee t and further indicates that dextromethorphan inhibits the serotonin uptake carrier at these doses, as a decrease in 5HIAA/serotonin ratio is well known to occur with serotonin uptake inhibitorst4. The mechanism for this decrease is thought to be a reduction in serotonin synthesis and turnover secondary to increased concentrations of serotonin in the synaptic cleft when the membrane uptake carrier is blocked. TABLE I
D~'tromethorphan lowers brain 5HI/IA concentration m rats Dextromethorphan was injected 140 min before rats were killed. Mean values+standard errors for 6 rats per group are shown. Asterisk indicate significant drug effect (P < 0.05).
Dose of dextrom,,thorphan (mg / kg, i.p)
5HIAA in whole brain (nmoles /g)
0 25 75
2.17±0.12 1 69±0.07 * (-22%) 1.64±0.15 * (-25%)
325
Various measures, including 5-hydroxytryptophan accumulation after decarboxylaxe inhibition and incorporation of radioactive tryptophan into 5-hydroxyindoles, show a reduction in serotonin synthesis when uptake is inhibited t4. As an electrophysiological correlate of these neurochemical measures, the firing of serotonin neurons in raphe areas is reduced in the presence of uptake inhibitors 5'23. Dextromethorphan is able to inhibit the serotonin uptake carrier in vivo because it has relatively high affinity for that uptake carrier. The IC5o concentration for dextromethorphan as an inhibitor of serotonin uptake by blood platelets in vitro is 120 nM t. In contrast, dextromethorphan inhibits calcium uptake in PC12 cells in vitro with an IC5o of I01 /zM 3, a concentration nearly 3 orders of magnitude higher. These considerations and our current findings suggest the most likely explanation for the dextromethorphan antagonism of serotonin neurotoxicity 10 days after PCA injection, as described by Finnegan et al. 7, is inhibition of the serotonin uptake carrier by dextromethorphan. Dextromethorphan has been reported to have neuroprotective effects in models of hypoxia-ischemia in laboratory animals in vivo 2z2~. There is no reason to suspect that inhibition of serotonin uptake is involved in those actions of dextromethorphan. Whether the effects of dextromethorphan in these models are due to antagonism of excitatory amino acid receptors, inhibition of calcium entry, actions on opioid receptors, or some other mechanism, the possibility must be considered that the same mechanisms account for its protection against PCA neurotoxicity, But because it is already well known that serotonin uptake inhibitors block the neurotoxic actions of PCA~'ts'2s, the protective effects of dextromethorphan do not necessarily implicate excitatory amino acid receptors or calcium entry in the neurotoxic actions of PCA. If agents that block calcium entry selectively were found to protect against PCA neurotoxicity, that could implicate altered calcium homeostasis in the neurotoxic actions of PCA. I Ahtee, L., Dextromethorphan inhibits 5.hydroxytryptamine uptake by human blood platelets and decreases 5-hydroxyindoleacetic acid content in rat brain, J. Pharm. Pharmacol., 27 (1975) 177-180. 2 Carlsson, A., Corrodi, H., Fuxe, K. and H0kfeit, T., Effect of antidepressant drugs on the depletion of intraneuronal brain 5-hydroxytryptamine stores caused by 4-methyl.a-ethyl-metatryamine, Ear. J. PharmacoL, 5 (1969) 357-366. 3 Carpenter, C.L., Marks, S.S., Watson, D.L. and Greenberg, D.A., Dextromethorphan and dextrorphan as calcium channel antagonists, Brain Res., 439 (1988) 372-375. 4 Choi, D.W., Dextrorphan and dextromethorphan attenuate glutamate neurotoxicity, Brain Res., 403 (1987) 333-336. 5 Clemens, J.A., Sawyer, B.D. and Cerimele, B., Further evidence
that serotonin i,~ a neurotransmitter involved in the control of prolactin secretion, Endocrinology, 100 (1977) 692-698. 6 Commins, D.L., Axt, K.J., Vosmer, G. and Seiden, L.S., Endogenously produced 5,6-dihydroxytryptamme may medmte the neurotoxic effects el para-chloroamphetamine, Brain Res., 419 (1987) 253-261. 7 Finnegan, K.T, Kerr, J.T. and Langston, J.W., Dextromethorphan protects against the neurotox~c effects of p-chloroamphetamine in rats, Brain Res., 558 (1991) 109-1il. 8 Fuller, R.W., Effects of p-chioroamphetamine on brain serotonin neurons, Neurochem. Res., 17 (1992)449-456. 9 Fuller, R.W., Mechanism by which uptake inhibitors antagomze p-chloroamphetamine-induced depletion of brain serotonin, Neurochem. Res., 5 (1980) 241-245. 10 Fuller, R.W. and Perry, K.W., Effects of buspirone and its metabolite, 1-(2-pyrimidinyl)piperazine,on brain monoammes and their metabolites w rats, J. Pharmacol. Exp. Ther., 248 (1989) 50-56. 11 Fuller, R.W., Perry, K.W. and Baker, J.C., Duration of the effects of a-ethyl-4-methyi-m-tyramine, (H75/12)on bram 5-hydroxyindole concentratiuns in rats, J. Pharm. PharmacoL, 28 (1976) 649-650. 12 Fuller, R.W., Perry, K.W. and Moiloy, B.B., Effect of 3-(p-trifluoro-methylphenoxy)-N-methyl-3-phenylpropylamine on the depletion of brain serotonin by 4-chloroamphetamine, J. Pharmacol. Exp. Ther,, 193 0975)796-803. 13 Fuller, R.W., Perry, K.W. and Molloy, B.B., Reversible and irreversible phases of serotonin depletion by 4-chloroamphetamine, Eur. J. PharmacoL, 33 (1975) 119-124. 14 Fuller, R.W. and Wong, D.T., Serotonin reuptake blockers in vitro and in rive, J. Chn. Psychopharmacol., 7 (1987) 36S-43S. 15 Harvey, J.A., McMaster, S.E. and Fuller, R.W., Comparison between the neurotoxic and serotonin-depleting effects of various halogenated derivatives of amphetamine in the rat, J. PharmacoL Exp. Ther., 202 (1977) 581-589. 16 Hashimoto, K. and Goromaru, T., High-affinity [~H]6-nitroquipazine binding sites in rat brain, Ear, J, Phamtacol., 180 (1990) 273-281. 17 Mamounas, L.A, and Molliver, M.E., Evidence for dual serotonergic projections to neocortex: axons from the dorsal and median raphe nuclei are differentially vulnerabl~ to the neurotoxin pchloroamphetamine (PCA), Exp. NearoL, 102 (1988) 23-36. 18 Meek, J.L., Fuxe, K. and Carlsson, A., Blockade of p.chloromethamphetamine induced 5-hydro~trypt:~mine depletion by chlorimipramine, chlorpheniramine and meperidine, Biochem. Pharmacol., 20 (I 971) 707-709. 19 Moffat, J,A. and Jhamandas, K., Effects of acute and chronic methadone treatment of the uptake of 31ol-5-hydroxytryplamine in rat hypoth:~lamu~:dices, Ear. J. Pharmacol., 36 (1976) 289-297. 20 Molliver, M.E., Berger, U.V., Mamounas, L.A., MoUiver, D.C., O'Hearn, E. and Wilson, M.A., Neurotoxicity of MDMA and related compounds: anatomic studies, Ann. N.Y. Acad. Sc'i., 600 (1990) 640-664. 21 Perrone, M.H., Luttinger, D., Hamel, L.T., Fritz, P.M., Ferraini, g. and Haubrich, D.R., In vivo assessment of napamezole, an alpha-2 adrenoceptor antagonist and monoamine re-uptake inhibitors, J. Pharmacol. Exp. Ther., 254 (1990) 476-483. 22 Prince, D.A. and Feeser, H.R., Dextromethorphan protects against cerebral infarction in a rat model of hypoxia-isehemia, Neurosci. Lett., 85 (1988) 291-296. 23 Rigdon, G.C. and Wang, C.M., Serotonin uptake blockers inhibit the firing of presumed serotonergic dorsal raphe neurons in vitro, Drag DeL'. Res., 22 (199t) 135-140. 24 Sanders-Bush, E., Bushing, J.A. and Sulser, F., Long-term effects of p-chloroamphetamine on tryptophan hydroxylase activity and on the !evels of 5.hydroxytryptamine and 5.hydro~indoleacetic acid in brain, Fur. J. Pharmacol., 20 (1972) 385-388. 25 Sanders-Bush, E. and Steranka, L.R., Immediate and long-term effects of p.chloroamphetamine on brain amines, Am~. N.Y. Acad. Sct., 305 (1978) 208-221.
326 26 Sinclair, J.G. and Lo, G.F., The blockade of serotonin uptake into synaptosomes: relationship to an interaction with monoamine oxidase inhibitors, Can. I. PhysioL Pharmacol., 55 (1977) 180-187. 27 Steinberg, G.K., Saleh, J. and Kunis, D., Delayed treatment with dextromethorphan and dextrorphan reduces cerebral damage after transient focal inschemia, Neurosct. Left, 89 (1988) 193-197.
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