Enhancement of [H]dopamine release and its [H]metabolites in rat striatum by nicotinic drugs

Recent evidence has identified directly muscarinic acetylcholine receptor (m-ACh R) and nicotinic acetylcholine receptor (n-ACh R) in the brain utilizing receptor binding assay. Several studies suggest that release of dopamine (DA) in the striatum is regulated by presynaptic receptors present on dopaminergic terminals. In the present study, the effects of cholinergic drugs on [³H]DA release were examined using micropunched tissue and synaptosomes obtained from rat striatum. ACh (5 × 10⁻⁴M) significantly increased spontaneous [³H]DA release, and the overflow was partially inhibited by d-tubocurarine (1 mM) but not atropine. Nicotine, lobeline, coniine and spartein, nicotinic agonists, significantly increased spontaneous and 25 mM K⁺ evoked [³H]DA release almost in a dose-dependent manner. In contrast, oxotremorine (2 × 10⁻⁴M), muscarinic agonist, did not any change in [³H]DA efflux. Furthermore, the metabolites of [³H]DA were separated by column chromatography. The main metabolite of [³H]DA in the spontaneous release from rat striatal synaptosomes was [³H]DOPAC (3,4-dihydroxyphenylacetic acid). Lobeline (5 × 10⁻⁵M) accelerated the outflow of [³H]DOPAC and [³H]OMDA metabolites (O-methylated and deaminated metabolites).These results could give rise to the suggestion that there was n-ACh R on the dopaminergic nerve terminals in the striatum and n-ACh R might have related to a directly excitatory effect on the DA release.     

Muscarinic receptors on dopamine terminals in the cat caudate nucleus: Neuromodulation of [H]dopamine release in vitro by endogenous acetylcholine

The directly acting muscarinic receptor agonist oxotremorine (1.8–10 μM) produced an increase in electrically evoked [³H]dopamine release from slices of the cat caudate. The maximal increase caused by oxotremorine was 40%, and was antagonized by the muscarinic receptor blocking agent atropine (0.1 μM). Exposure to the acetylcholinesterase (AChE) inhibitor physostigmine (1 μM) resulted in a 50% increase in electrically evoked [³H]dopamine release. The increase caused by physostigmine was also antagonized by atropine (0.1 μM).Atropine did not, however, alter the modulations in [³H]dopamine release mediated by the dopamine autoreceptor: the increase in electrically evoked [³H]dopamine release caused by the dopamine receptor antagonist S-sulpiride (0.1 μM) and the decrease caused by the dopamine receptor agonist pergolide (30 nM) were unaffected by atropine (0.1 μM). These results indicate that the muscarinic receptor-mediated and dopamine autoreceptor-mediated presynaptic effects on [³H]dopamine release are independent.The present results suggest that in the electrically depolarized caudate slice in vitro, released endogenous acetylcholine may interact with muscarinic receptors faciliting depolarization-evoked [³H]dopamine release,ifAChE is inhibited. These muscarinic receptors may be located on dopamine nerve terminals. In the context of present neuroanatomical knowledge, the action of released endogenous acetylcholine on dopamine terminals may be a non-synaptic neuromodulation.     

Is the muscarinic receptor that mediates potentiation of dopamine release negatively coupled to the cyclic GMP system?

Dopamine (DA) terminals in rat corpus striatum and frontal cortex possess muscarinic receptors that mediate enhancement of the depolarization-evoked release of the catecholamine. The effects of the membrane-permeating cyclic guanosine monophosphate (cyclic GMP) analog 8-Br-cyclic GMP and of the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX) on the muscarinic-induced increase of DA release were investigated in striatal synaptosomes prelabeled with [3H]DA and exposed in superfusion to 15 mM KCl and to acetylcholine (ACh). Preincubation of synaptosomes with 8-Br-cyclic GMP (10-200 microM) or with IBMX (200 microM) prevented the ACh-induced enhancement of [3H]DA release, without affecting the K+-evoked release of the [3H]amine. No significant decrease of the ACh effect was observed when 8-Br-cyclic GMP or IBMX were added concomitantly with ACh to the superfusion medium. The data suggest that stimulation of presynaptic muscarinic receptors on DA terminals may produce enhancement of 3H DA release through a decrease of the intraterminal cyclic GMP content.     

Presynaptic modulation of the release of dopamine from striatal synaptosomes: Differences in the effects of high K stimulation, methamphetamine and nicotinic drugs

We examined the effects of nicotinic drugs on the metabolites of [3H]dopamine (DA) released from rat striatal synaptosomes in comparison with the effects of high K+ and methamphetamine stimulation. The pattern of outflow of [3H]DA and its metabolites induced by nicotine stimulants, acetylcholine (ACh), nicotine and lobeline, was different from that induced by high K+ and methamphetamine.     

Muscarinic receptors on dopamine terminals in the cat caudate nucleus: neuromodulation of [3H]dopamine release in vitro by endogenous acetylcholine

The directly acting muscarinic receptor agonist oxotremorine (1.8-10 microM) produced an increase in electrically evoked [3H] dopamine release from slices of the cat caudate. The maximal increase caused by oxotremorine was 40%, and was antagonized by the muscarinic receptor blocking agent atropine (0.1 microM). Exposure to the acetylcholinesterase (AChe) inhibitor physostigmine (1 microM) resulted in a 50% increase in electrically evoked [3H]dopamine release. The increase caused by physostigmine was also antagonized by atropine (0.1 microM). Atropine did not, however, alter the modulations in [3H]dopamine release mediated by the dopamine autoreceptor: the increase in electrically evoked [3H]dopamine release caused by the dopamine receptor antagonist S-sulpiride (0.1 microM) and the decrease caused by the dopamine receptor agonist pergolide (30 nM) were unaffected by atropine (0.1 microM). These results indicate that the muscarinic receptor-mediated and dopamine autoreceptor-mediated presynaptic effects on [3H]dopamine release are independent. The present results suggest that in the electrically depolarized caudate slice in vitro, released endogenous acetylcholine may interact with muscarinic receptors facilitating depolarization-evoked [3H]dopamine release, if AChE is inhibited. These muscarinic receptors may be located on dopamine nerve terminals. In the context of present neuroanatomical knowledge, the action of released endogenous acetylcholine on dopamine terminals may be a non-synaptic neuromodulation.     

Angiotensin II inhibits the release of [3H]acetylcholine from rat entorhinal cortex in vitro

The effects of angiotensin I and II on basal potassium-induced release of [3H]acetylcholine were investigated in slices of rat entorhinal cortex. Potassium (10-25 mM) produced a concentration-dependent increase in the release of [3H]acetylcholine in the presence of extracellular calcium. Angiotensin II (10(-9)-10(-5) M) (but not angiotensin I) reduced the potassium-induced release of [3H]acetylcholine in a concentration-related manner to 60% of control levels, but did not effect basal tritium release. The effect of angiotensin II was antagonised by [1-sarcosine, 8-threonine] angiotensin II, an angiotensin II receptor antagonist, but not by agents acting on alpha- and beta-adrenoceptors, muscarinic, nicotinic, histamine or 5-hydroxytryptamine receptors nor by the angiotensin converting enzyme (ACE) inhibitor SQ 29852. The results indicate that angiotensin II acting via an angiotensin II receptor can inhibit the release of [3H]acetylcholine in slices of the rat entorhinal cortex. It is hypothesised that the ability of ACE inhibitors to facilitate cognitive processes may be related to a reduced availability of angiotensin II.     

Biochemical evidence for the presence of presynaptic receptors on dopaminergic nerve terminals

Two compartments of striatal synaptosome dopamine were identified by differential labelling with the isotopic presursors, L-tyrosine and Dopa, and from specific radioactivity measurement. Either, endogenous or exogenous L-tyrosine could provide a source for the dopamine pool synthesised and released in response to K+ depolarization, whereas external DOPA did not enter this pool. Acetylcholine (0.1 mM) in the presence of neostigmine (0.1 mg/ml) increased dopamine turnover as shown by increased formation of [14C] dopamine and [14C] DOPAC from [14C] DOPA. Haloperidol (0.65 mM) did not affect the size of dopamine pools but increased the conversion of [14C] DOPA to [14C] dopamine and the formation of [14C] DOPAC. Acetylcholine stimulated the release of dopamine from synaptosomes, which effect could be modified by both muscarinic and nicotinic antagonists. In the presence of the muscarinic antagonist, atropine, acetylcholine stimulated dopamine release, whereas in the presence of the nicotinic antagonists, hexamethonium (0.2 mM) or alpha-bungaro-toxin (0.188 muM), acetylcholine inhibited dopamine release. This showed that presynaptic cholinergic receptors were operational, excitatory nicotinic receptors in the former case and inhibitory muscarinic in the latter. The nicotinic receptor was shown to be saturable and to bind specifically 11.2 fmoles of [3H] alpha-bungarotoxin per mg. protein which could be prevented by hexamethonium or D-tubocurarine.     

Pharmacological characterization of adrenal paraneurons: substance P and somatostatin as inhibitory modulators of the nicotinic response

A pharmacological study was made of the effects of various muscarinic and nicotinic agonists and their antagonists on the release of [3H]noradrenaline ([3H]NA) from cultures of isolated bovine adrenal medullary cells. A study was also made of the effects of substance P and somatostatin on the release of [3H]NA evoked by nicotinic agonists. By 2 days in culture these adrenal 'paraneurons' had developed long varicose processes with growth cones and generally resembled noradrenergic neurons in culture. In the present study, adrenal paraneurons were incubated with [3H]NA which was taken up and stored in reserpine-sensitive sites. Exposure of the cultures to acetylcholine (ACh) resulted in release of [3H]NA into the external medium. High concentrations of K+ (56 mM) also evoked release of [3H]NA. The release of [3H]NA induced by ACh or K+ (56 mM) was Ca2+-dependent. Pharmacological studies with nicotinic (ACh, nicotine) and muscarinic (methacholine, pilocarpine) agonists and their antagonists (mecamylasmine, d-tubocurarine, hexamethonium; and atropine, scopolamine, respectively) showed that the adrenal paraneurons contained only nicotinic receptors. Substance P produced a dose-dependent inhibition of ACh (5 x 10(-5) M) stimulated [3H]NA release in the range of 10(-8) to 5 x 10(-5) M with an ID50 of 10(-6) M. A similar inhibition of NA release by substance P was obtained when nicotine (K X 10(-6) M) was used as the agonist, but not when K+ (50 MM) was used to depolarize the cells. Substance P (10-10) to 5 x 10(-5) M) by itself did not have a significant effect on the basal release rate of [3H]NA from these cells. Somatostatin at relatively high concentrations (10(-6)-10(-3) M; ID50 2 x 10(-5) M) inhibited the release induced by ACh, but not by K+ (56 mM). The present results provide the first direct evidence at a cellular level that substance P and somatostatin act as inhibitory modulators of the nicotinic ACh response, and support a role for these peptides as inhibitory neuromodulators at nicotinic receptor sites in the nervous system.     

Kinetics of dopamine and noradrenaline transport in synaptosomes from cerebellum, striatum and frontal cortex of normal and reeler mice.

Recent evidence indicates that the cerebellum has a dopaminergic system. In order to elucidate further the dopaminergic system in the cerebellum, we investigated the transport of dopamine (DA) in synaptosomal preparations of normal and reeler mice. For comparative purposes we also studied DA transport in synaptosomal preparations from striatum and frontal cortex and compared DA transport to noradrenaline (NA) transport. [3H]-DA transport into cerebellar synaptosomes was found to be a Na(+)-dependent, two component system--a high affinity, low capacity and a low affinity, high capacity. In striatum [3H]-DA is transported by a similar high but different low affinity component. Maximal velocities of both transport components in the striatum were higher than the corresponding ones in the cerebellum. In the frontal cortex we also observed two [3H]-DA transport components with affinities significantly lower than those in cerebellum and striatum. [3H]-NA transport into synaptosomes, prepared from the three brain regions studied, showed two transport components with similar Kt and Vmax values, except for the high affinity component in striatum whose affinity is lower. In reeler mice [3H]-DA transport was different from normal only in the cerebellum where the maximal velocity for both transport components was significantly higher (2x). In contrast, no significant difference was observed in the transport of [3H]-NA. The accumulated [3H]-DA from cerebellar slices was found to be releasable by K+ stimulation, in a Ca(++)-dependent manner, and most of the released radioactivity was in the form of [3H]-DA. These results indicate that in the cerebellum there is a low-density dopaminergic system which is distinct from the corresponding noradrenergic system.     

Characterization of the excitatory amino acid receptor-mediated release of [H]acetylcholine from rat striatal slices

The pharmacological nature of the interaction of excitatory amino acids with striatal cholinergic neurons was investigated in vitro. Agonists of excitatory amino acid receptors evoked the release of [³H]acetylcholine from slices of rat striatum, in the presence of magnesium (1.2 mM). Removal of magnesium from the medium markedly increased the release of [³H]acetylcholine evoked by all excitatory amino acid receptor agonists tested, with the exception of kainate. In the absence but not the presence of magnesium, a clear rank order of potency was found: N-methyl-dl-aspartate = ibotenate >l-glutamate >l-aspartate cysteate > kainate = quisqualate.The excitatory amino acid receptor mediating [³H]acetylcholine release resembles the N-methyl-d-aspartate preferring (N-type) receptor, as previously characterized electrophysiologically, according to 3 criteria: (1) rank order of potency of agonists; (2) magnesium-sensitivity; and (3) antagonism by 2-amino-5-phosphonovalerate.The release of [³H]acetylcholine evoked by N-methyl-dl-aspartate was blocked by tetrodotoxin (0.5 μM). Moreover, N-methyl-dl-aspartate failed to evoke [³H]acetylcholine release from slices of hippocampus, where cholinergic afferents, rather than interneurons, are found. These results suggest that excitatory amino acids act at receptors on the dendrites of striatal cholinergic interneurons, giving rise to action potentials and release of acetylcholine from cholinergic nerve terminals.     

Comparative evaluation of [3H]WIN 35428 and [3H]GBR 12935 as markers of dopamine innervation density in brain

WIN 35428 and GBR 12935, two uptake blocker ligands of the membrane transporter for dopamine (DA), were evaluated as quantitative markers of DA innervation density in CNS tissue. From alternate rat brain slices respectively processed for either light microscope or film autoradiography, counts of DA axon terminals (varicosities) labeled by uptake/storage of [³H]DA were matched with densitometric measurements of the specific binding of [³H]WIN 35428 and [³H]GBR 12935 in the same anatomical areas. The relation between the two parameters was examined in 1) the normal cingulate cortex; 2) the neostriatum severely DA-denervated by unilateral intramesencephalic injections of 6-hydroxydopamine; and 3) the neostriatum, partly DA-reinnervated by an intrastriatal graft of fetal mesencephalic neurons after prior 6-hydroxydopamine lesion. For technical reasons, the hyperdense DA innervation of normal striatum was not amenable to such correlative testing. Data were subjected to multilevel analysis. Specific [³H]WIN binding at 37°C was tightly and linearly correlated with the number of DA varicosities over the full range of DA innervation densities tested. The regression lines for intact cortex and for DA-denervated as well as DA-reinnervated neostriatum had the same slope and crossed the ordinate near zero. In contrast, [³H]GBR 12935 binding at 37°C showed no correlation with the number of DA varicosities. A linear correlation could be obtained after incubation with [³H]GBR 12935 at 4°C in the presence of ZnSO₄, but the intercept of this regression line remained significantly above zero at origin, indicating extraneous binding to non-DA transporter sites. Providing that the hyperdense DA innervation of the normal neostriatum does not generate a particular problem in vivo as it does in vitro, WIN 35428, but not GBR 12935, might satisfy the selectivity and sensitivity requirements of a quantitative marker of DA innervation density for eventual use in positron emission tomographic studies. Synapse 25:163–175, 1997. © 1997 Wiley-Liss, Inc.     

Muscarinic receptor subtypes modulate the release of [3H]-noradrenaline in rat spinal cord slices

Spinal muscarinic receptors are involved in the mediation of antinociceptive effects. The modulation of noradrenaline (NA) release on muscarinic receptor subtypes in the rat spinal cord was investigated in in vitro perfusion experiments. After rat spinal cord slices were preincubated in [3H]NA, the slices were perfused with a superfusion apparatus. The slices were field stimulated during the 4th (S1) and 11th (S2) superfusion collection periods. Perfusion of drugs was initiated at the 8th collection period and was maintained until the 14th collection period. Fractional release was calculated as the percentage of the radioactivity present in the slices at the beginning of the stimulation period. Drugs were administered between S1 and S2. The following drugs were used: [3H]NA, neostigmine, pirenzepine (M1 antagonist), AFDX116 (M2 antagonist), atropine. Neostigmine significantly increased the release of [3H]NA in a concentration-dependent manner. Pirenzepine (1 microM) and atropine (0.3 microM) significantly reduced the release of [3H]NA, but AFDX116 (1 microM) did not significantly reduce release in the presence of neostigmine (1 microM). The results of this study indicate that neostigmine can enhance noradrenergic neurotransmission, and that acetylcholine can stimulate spinal cord NA release via M1 muscarinic receptor subtypes.     

Pharmacological and mechanistic studies of cholecystokinin-facilitated [3H]dopamine efflux from rat nucleus accumbens

Cholecystokinin (CCK) is co-localized with dopamine (DA) in mesolimbic neurons of the CNS and appears to selectively regulate the output of this system. In an attempt to characterize the nature of CCK interactions with mesolimbic DA-containing nerve terminals, we have investigated CCK regulation of [3H]DA overflow from rat nucleus accumbens slices. CCK-8 produced a saturable and potent (EC50 = 3 nM) facilitation of KCl (35 mM)-evoked [3H]DA efflux from nucleus accumbens, but failed to significantly alter [3H]DA efflux from striatum: The stimulatory action of CCK-8 was unaffected by the muscarinic antagonist atropine, the opiate antagonist naloxone, or the selective ion channel blockers tetrodotoxin and nifedipine. Pharmacological studies revealed that non-sulfated CCK-8 and CCK-4 (up to low micromolar concentrations) did not facilitate [3H]DA efflux from accumbens slices. Furthermore, the effect of CCK-8 was selectively and potently (IC50 = 300 nM) inhibited by the Type A-selective CCK antagonist CR-1409. Taken together, these results indicate that CCK regulates DA efflux from mesolimbic nerve terminals via a direct presynaptic action on receptors which display a pharmacological profile that is similar to Type A CCK receptors in gastrointestinal tissues.     

Somatostatin and cholecystokinin octapeptide differentially modulate the release of [H]acetylcholine from caudate nucleus but not cerebral cortex: Role of dopamine receptor activation

The effects of somatostatin (SOM) and cholecystokinin octapeptide (CCK-8) on basal and potassium-induced release of acetylcholine (ACh) were investigated in slices of rat caudate nucleus (CN) and, for comparison, cerebral cortex (CX). Potassium (5–55 mM) produced a concentration-dependent increase in the release of [³H]ACh in the presence of extracellular Ca²⁺. SOM (1 μ), CCK-8 (1 μM) and the dopamine (DA) receptor agonist, apomorphine (APO, 30 μM) inhibited the K⁺-induced (35 mM) release of [³H]ACh by 26–32% from CN, but did not affect ACh release from CX. Other peptides (1 μM), such as Met-enkephalin, vasoactive intestinal peptide, thyrotropin-releasing hormone and substance P, had no effect on release of [³H]ACh in CN or CX. Sulpiride (SULP), a dopamine receptor antagonist, prevented the effects of APO and SOM, but not CCK-8, to inhibit [³H]ACh release. The results indicate that: (1) SOM and CCK-8 inhibit the release of [³H]ACh in CN, but not CX; and (2) the inhibitory effect of SOM, but not CCK-8, on [³H]ACh release is mediated by dopaminergic mechanisms.     

Evidence that ibogaine releases dopamine from the cytoplasmic pool in isolated mouse striatum

Summary We measured the effect of ibogaine on the tritium efflux from isolated mouse striatum preloaded with [³H]dopamine ([³H]DA). Ibogaine increased the basal tritium outflow in a concentration-dependent manner, but it was without effect on electrical stimulation-induced tritium overflow. Separation of the released radioactivity after ibogaine administration showed that this drug increased the release of [³H]DA and [³H]-dihydroxyphenylacetic acid ([³H]DOPAC), but the efflux of O-methylated-deaminated metabolites was not changed. The dopamine (DA)-releasing effect of ibogaine was reduced by the DA uptake inhibitors cocaine and nomifensine. The tritium efflux evoked by ibogaine was not altered by omission of Ca²⁺ from the perfusion buffer or by inhibition of the voltage-sensitive Na⁺ channels with tetrodotoxin. Ibogaine maintained its effect on release from superfused striatum prepared from reserpine-pretreated mice. The ibogaine-induced tritium release measured from mouse striatum that was preloaded with [³H]DA was not affected by the D-2 DA receptor ligands (–)-quinpirole and (+/–)-sulpiride, indicating that the ibogaine-induced release is not subject to presynaptic autoreceptor regulation. Ibogaine failed to affect [³H]DA uptake and retention in mouse striatum. These data indicate that at the nerve terminal level ibogaine releases DA, and the primary source for the release is probably the cytoplasmic pool. The DA-releasing effect of ibogaine may have importance in mediation of its hallucinogenic action, as seen in a frequent practice in African cults.     

High-affinity binding of [3H]acetylcholine to muscarinic cholinergic receptors

High-affinity binding of [3H]acetylcholine to muscarinic cholinergic sites in rat CNS and peripheral tissues was measured in the presence of cytisin, which occupies nicotinic cholinergic receptors. The muscarinic sites were characterized with regard to binding kinetics, pharmacology, anatomical distribution, and regulation by guanyl nucleotides. These binding sites have characteristics of high-affinity muscarinic cholinergic receptors with a Kd of approximately 30 nM. Most of the muscarinic agonist and antagonist drugs tested have high affinity for the [3H]acetylcholine binding site, but pirenzepine, an antagonist which is selective for M-1 receptors, has relatively low affinity. The ratio of high-affinity [3H]acetylcholine binding sites to total muscarinic binding sites labeled by [3H]quinuclidinyl benzilate varies from 9 to 90% in different tissues, with the highest ratios in the pons, medulla, and heart atrium. In the presence of guanyl nucleotides, [3H] acetylcholine binding is decreased, but the extent of decrease varies from 40 to 90% in different tissues, with the largest decreases being found in the pons, medulla, cerebellum, and heart atrium. The results indicate that [3H]acetylcholine binds to high-affinity M-1 and M-2 muscarinic receptors, and they suggest that most M-2 sites have high affinity for acetylcholine but that only a small fraction of M-1 sites have such high affinity.     

Acetorphan,an Enkephalinase Inhibitor,Modulates Dopaminergic Transmission in Rat Olfactory Tubercle,but not in the Nucleus Accumbens and Striatum

The present study focused on the effects of acetorphan, a parenterally active enkephalinase inhibitor, on dopaminergic transmission in rat olfactory tubercle, nucleus accumbens and striatum. Acetorphan was administered i.v. (10 mg/kg) 15 min before measurement of the in vivo specific binding of [3H]N-propylnorapomorphine ([3H]NPA) or measurement of the levels of dopamine (DA) and its metabolites 3-methoxytyramine-homovanillic acid (3MT-HVA) in the three areas. Acetorphan decreased the in vivo specific binding of [3H]NPA in the olfactory tubercle, this effect being antagonized by naloxone 1.5 mg/kg s.c. DA release in this brain structure was also significantly increased by acetorphan 10 mg/kg, as indicated by the 3MT:DA and HVA:DA ratios. Neither the specific binding of [3H]NPA nor DA metabolism and release were modified by the inhibitor in the striatum and the nucleus accumbens. The stimulant effect of acetorphan was significantly decreased in rats in which a bilateral lesion of dopaminergic endings in the olfactory tubercle had been produced by 6-hydroxydopamine (6-OHDA). These results suggest that dopaminergic transmissions in the olfactory tubercle are particularly sensitive to the modulation exerted by endogenous enkephalins, this modulation being at least partly involved in the increased locomotion induced by the enkephalinase inhibitor.     

Effect of chronic lithium on cholinergically mediated responses and [3H]QNB binding in rat brain

Lithium (Li) has been previously reported to increase acetylcholine turnover and release in rat brain and to potentiate the neurotoxicity of cholinergic agents. We studied the effect of chronic Li administration, alone and in combination with the muscarinic antagonist, scopolamine, on two cholinergically-mediated responses and on muscarinic cholinergic receptor (MCR) binding in rat brain. Administered separately, Li and scopolamine enhanced the cataleptic and hypothermic responses to pilocarpine; combined administration resulted in an additive effect on both these measures. [3H]Quinuclidinyl benzilate ([3H]QNB) binding was increased by Li in the corpus striatum but not in the cortex, hippocampus and hypothalamus. Scopolamine increased [3H]QNB binding in the striatum, cortex and hippocampus; Li and scopolamine effects on striatal MCR were not additive. Contrary to a previous report, antagonist-induced MCR supersensitivity was not prevented by concurrent Li administration in any of the brain areas studied. The additive effect of Li and scopolamine on pilocarpine-induced catalepsy and a trend in this direction for pilocarpine-induced hypothermia suggest that the actions of the two agents to enhance cholinergically mediated responses may be achieved by different mechanisms. Supersensitive responses following scopolamine may be attributed to antagonist-induced up-regulation of postsynaptic muscarinic receptors as demonstrated in the binding studies. The effects of Li to enhance cholinergically-mediated catalepsy and hypothermia are interpreted as extending previous reports that Li stimulates brain cholinergic function by a presynaptic increase in acetylcholine turnover and release.     

Effects of verapamil on the release of different neurotransmitters

The effect of verapamil on resting and depolarization-induced monoamine release was investigated in rat hippocampal synaptosomes prelabeled with [³H]-5-hydroxytryptamine (HT) or [³H]-norepinephrine (NE) and rat striatal synaptosomes prelabeled with [³H]-dopamine (DA). Verapamil (50 μM) completely abolishes high K⁺-induced [³H]-NE release, but paradoxically facilitates high K⁺-induced [³H]-5-HT and [³H]-DA release. All these high K⁺-evoked responses were Ca²⁺ dependent. Verapamil does not modify [³H]-NE baseline release, but increases dose dependently [³H]-5-HT and [³H]-DA baseline release. Verapamil (10 μM, for 5 min) increases endogenous DA release (70%) and endogenous 5-HT release (40%) independently on the presence of external Ca²⁺. The total amount of these monoamines (released plus retained by the preparation) and their metabolites (DOPAC and 5-HIAA) was similar in control and verapamil-treated synaptosomes. Verapamil displaces [³H]-spiroperidol specific binding (K_i of 2.4 × 10^?6M) and [³H]-SCH-23390 specific binding (K_i of 9 × 10^?6M) from striatal synaptosomal membranes, and [³H]-5-HT specific binding (K_i of 3 × 10^?5M) from hippocampal synaptosomal membranes. It is concluded that in addition to the Ca²⁺ antagonistic properties of verapamil on the Ca²⁺-dependent, depolarization-induced release of some neurotransmitters [gamma aminobutyric acid (GABA and NE)], another mechanism probably mediated by presynaptic receptors underlies the effects of verapamil on DA and 5-HT release from discrete brain regions. © 1995 Wiley-Liss, Inc.     

Nitric oxide induces calcium-dependent release of [3H]dopamine release from striatal slices

Hydroxylamine (0.01–30 mM), a nitric oxide (NO) generator, produced a concentration-dependent release of [³H]dopamine ([³H]DA) from rat striatal slices. Hemoglobin (10 μM), a NO scavenger, reduced basal [³H]DA release and blocked hydroxylamine (100 μM)-stimulated [³H]DA efflux. Tetrodotoxin (0.5 μM) had no significant effect. Sodium cyanide was used as a model compound to test the possibility that NO acted through blockade of mitochondrial electron transport. Calcium-free experimental buffer (1 mM EGTA) reduced basal release and the hydroxylamine response, while sodium cyanide-induced release did not change under these experimental conditions. Cadmium (200 μM), a non-selective inhibitor of voltage-dependent calcium channels, reduced the hydroxylamine response by 69%. Methylene blue (10 μM), an inhibitor of guanylate cyclase, produced a 3-fold increase in the basal release but had no significant effect on the hydroxylamine response. These data suggest that NO induces calcium-dependent [³H]DA release from the striatum via a mechanism which is independent of blockade of electron transport or activation of guanylate cyclase. © 1993 Wiley-Liss, Inc.