Differential regulation of somatodendritic and nerve terminal dopamine release by serotonergic innervation of substantia nigra

Nigrostriatal dopaminergic neurons release dopamine from dendrites in substantia nigra and axon terminals in striatum. The cellular mechanisms for somatodendritic and axonal dopamine release are similar, but somatodendritic and nerve terminal dopamine release may not always occur in parallel. The current studies used in vivo microdialysis to simultaneously measure changes in dendritic and nerve terminal dopamine efflux in substantia nigra and ipsilateral striatum respectively, following intranigral application of various drugs by reverse dialysis through the nigral probe. The serotonin releasers (+/-)-fenfluramine (100 micro m) and (+)-fenfluramine (100 micro m) significantly increased dendritic dopamine efflux without affecting extracellular dopamine in striatum. The non-selective serotonin receptor agonist 1-(m-chlorophenyl)-piperazine (100 micro m) elicited a similar pattern of dopamine release in substantia nigra and striatum. NMDA (33 micro m) produced an increase in nigral dopamine of a similar magnitude to mCPP or either fenfluramine drug. However, NMDA also induced a concurrent increase in striatal dopamine. The D2 agonist quinpirole (100 micro m) had a parallel inhibitory effect on dopamine release from dendritic and terminal sites as well. Taken together, these data suggest that serotonergic afferents to substantia nigra may evoke dendritic dopamine release through a mechanism that is uncoupled from the impulse-dependent control of nerve terminal dopamine release.     

Inhibition of Ischemia-Induced Dopamine Release by ω-Conotoxin, a Calcium Channel Blocker, in the Striatum of Spontaneously Hypertensive Rats: In Vivo Brain Dialysis Study

The effect of omega-conotoxin GVIA (CgTX), an N-and L-type voltage-sensitive calcium channel (VSCC) blocker, on the release of dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) in the striatum before and during transient cerebral ischemia in spontaneously hypertensive rats was studied using an in vivo brain dialysis technique. Continuous perfusion of CgTX in the striatum was started 20 min before ischemia and concentrations of dopamine and DOPAC in the dialysate were measured using HPLC with an electro-chemical detector. Before ischemia, both 10 and 100 microM CgTX significantly lowered the concentration of dopamine, to 49% of the basal values. DOPAC concentrations also decreased significantly, by 28 and 17%, respectively. Forebrain ischemia, produced by bilateral carotid artery occlusion, reduced striatal blood flow to less than 6% of the resting value in each group. During 20 min of ischemia, the vehicle group showed a marked increase in dopamine (175 times the basal concentration). In the 10 or 100 microM CgTX perfusion group, in contrast, dopamine release was significantly attenuated, to 38 or 29% of the vehicle group, respectively. DOPAC concentrations decreased during ischemia to 58% of the basal value in the vehicle group and 49% in both CgTX groups. These results indicate that the massive release of striatal dopamine during ischemia depends largely on the influx of extracellular calcium via CgTX-sensitive VSCCs.     

Inhibition of Na+,K+-ATPase by Ouabain Opens Calcium Channels Coupled to Acetylcholine Release in Guinea Pig Myenteric Plexus

Abstract: Ouabain, an Na⁺,K⁺-ATPase inhibitor, increases the release of acetylcholine (ACh) from various preparations in a Ca²⁺-independent way. However, in other preparations the release of ACh evoked by ouabain is dependent on the presence of extracellular calcium. In the present study, we have labeled the ACh of myenteric plexus longitudinal muscles of guinea pig ileum and compared the effect of calcium channel blockers on ouabain-evoked release of [³H]ACh. Release of [³H]ACh evoked by ouabain is dose dependent and decreased markedly in the absence of calcium or in the presence of cadmium, a nonspecific calcium channel blocker. N-type calcium channel blockage by the ω-conotoxins GVIA (selective N-type calcium channel blocker) and MVIIC (a nonselective calcium channel blocker) inhibited by 45 and 55%, respectively, the release of [³H]ACh. L-type calcium channel suppression by low concentrations of verapamil, nifedipine, and diltiazem had no effect on the release of [³H]ACh. The release of transmitter was also not affected significantly by nickel, a T-type calcium channel blocker. In addition, ω-agatoxin-IVA, at concentrations that block P- and Q-type calcium channels, did not affect significantly the release of [³H]ACh. Thus, extracellular Ca²⁺ is essential for the release of ACh induced by ouabain from guinea pig ileum myenteric plexus. In this preparation, the N-type calcium channel plays a dominant role in transmitter release evoked by inhibition of Na⁺,K⁺-ATPase, but other routes of calcium entry in addition to these channels can also support the release of neurotransmitter induced by ouabain.     

Biochemistry of Somatodendritic Dopamine Release in Substantia Nigra: An In Vivo Comparison with Striatal Dopamine Release

Abstract: The somatodendritic release of dopamine in substantia nigra previously has been suggested to be nonvesicular in nature and thus to differ from the classical, exocytotic release of dopamine described for the dopaminergic nerve terminal in striatum. We have compared the effects of reserpine, a compound that disrupts vesicular sequestration of monoamines, on the storage and release of dopamine in substantia nigra and striatum of rats. Reserpine administration (5 mg/kg, i.p.) significantly decreased the tissue level of dopamine in substantia nigra pars reticulata, substantia nigra pars compacta, and striatum. In these brain areas, reserpine-induced reductions in tissue dopamine level occurred within 2 h and persisted at 24 h postdrug. In vivo measurements using microdialysis revealed that reserpine administration rapidly decreased the extracellular dopamine concentration to nondetectable levels in substantia nigra as well as in striatum. In both structures, it was observed that reserpine treatment significantly attenuated the release of dopamine evoked by a high dose of amphetamine (10 mg/kg, i.p.) given 2 h later. In contrast, dopamine efflux in response to a low dose of amphetamine (2 mg/kg, i.p.) was not altered by reserpine pretreatment either in substantia nigra or in striatum. The present data suggest the existence, both at the somatodendritic and at the nerve terminal level, of a vesicular pool of dopamine that is the primary site of transmitter storage and that can be displaced by high but not low doses of amphetamine. The physiological release of dopamine in substantia nigra and in striatum is dependent on the integrity of this vesicular store.     

Different effects of L-, N- and T-type calcium channel blockers on striatal dopamine release measured by microdialysis in freely moving rats.

Using a microdialysis method, we have investigated effects of the voltage-dependent calcium channel blockers, verapamil, nicardipine, omega-conotoxin and flunarizine on the dopamine release and metabolism in the striatum of freely moving rat. Perfusion of verapamil (1-300 microM) and nicardipine (1-100 microM), an L-type calcium channel blocker, into the striatum through the dialysis membrane showed a dose-dependent decrease of dopamine release in the dialysate and slight increase of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) levels. Treatment of omega-conotoxin (0.1, 1 microM), an N-type channel blocker, decreased about 50% basal dopamine release and slightly decreased DOPAC and HVA levels. Treatment with flunarizine (10 microM), an T-type channel blocker, did not affect the dopamine release and metabolism. From these data, it appears that treatments of the L- and N-type voltage-dependent calcium channel blockers in rat striatum suppress basal dopamine release, but T-type blocker does not suppress it, suggesting that L-, N- and T-type calcium channels regulate in vivo dopamine release in a different mechanism.     

Involvement of Calcium Channels in Depolarization-Evoked Release of Adenosine from Spinal Cord Synaptosomes

Abstract: The potential involvement of L- and N-type voltage-sensitive calcium (Ca²⁺) channels and a voltage-independent receptor-operated Ca²⁺ channel in the release of adenosine from dorsal spinal cord synaptosomes induced by depolarization with K⁺ and capsaicin was examined. Bay K 8644 (10 n M ) augmented release of adenosine in the presence of a partial depolarization with K⁺ (addition of 6 m M ) but not capsaicin (1 and 10 μ M ). This augmentation was dose dependent from 1 to 10 n M and was followed by inhibition of release from 30 to 100 n M . Nifedipine and nitrendipine inhibited the augmenting effect of Bay K 8644 in a dose-dependent manner, but neither antagonist had any effect on release of adenosine produced by K⁺ (24 m M ) or capsaicin (1 and 10 μ M ) ω-Conotoxin inhibited K⁺-evoked release of adenosine in a dose-dependent manner but had no effect on capsaicin-evoked release. Ruthenium red blocked capsaicin-induced release of adenosine but had no effect on K⁺-evoked release. Although L-type voltage-sensitive Ca²⁺ channels can modulate release of adenosine when synaptosomes are partially depolarized with K⁺, N-type voltage-sensitive Ca²⁺ channels are primarily involved in K⁺-evoked release of adenosine. Capsaicin-evoked release of adenosine does not involve either L- or N-type Ca²⁺ channels, but is dependent on a mechanism that is sensitive to ruthenium red.     

Effect of Voltage-Sensitive Calcium Channel Antagonists on the Release of 5-Hydroxytryptamine from Rat Hippocampus In Vivo

The effect of calcium channel antagonists on the release of 5-hydroxytryptamine from the hippocampus of the chloral hydrate-anaesthetised rat was studied using the technique of intracerebral microdialysis. As the basal concentration of 5-hydroxytryptamine was close to the limit of detection of the HPLC method (8 fmol), the 5-hydroxytryptamine reuptake inhibitor, fluoxetine (10 microM), was included in the perfusion fluid. The L-type voltage-sensitive calcium channel antagonists, PN200-110, diltiazem, and verapamil, all passed through the dialysis membrane, giving a recovery of 20-30%. The N-type voltage-sensitive calcium channel antagonist, omega-conotoxin, penetrated less readily (12% recovery). The dihydropyridine, PN200-110, adhered to the probe, resulting in an effective concentration at the membrane 30% of that in the perfusion fluid. The concentration of 5-hydroxytryptamine in the dialysate samples was reduced by 60% in the absence of calcium. The L channel antagonists had little effect on the release of 5-hydroxytryptamine, which was inhibited, in a dose-dependent manner, to a maximum of 40% by omega-conotoxin. It is concluded that, under physiological conditions, the release of 5-hydroxytryptamine from the rat hippocampus is dependent on the entry of calcium through N-type voltage-sensitive calcium channels, although another calcium channel may also be involved.     

Involvement of Different Types of Voltage-Sensitive Calcium Channels in the Presynaptic Regulation of Noradrenaline Release in Rat Brain Cortex and Hippocampus

Abstract: Transmitter release at the nerve terminal is mediated by the influx of Ca²⁺ through voltage-sensitive calcium channels (VSCCs). Many types of VSCCs have been found in neurons (T, N, L, and P), but uncertainty remains about which ones are involved in neuronal excitation-secretion coupling. Specific ligands for the L- and N-type VSCCs were used to determine which of these subtypes might be involved in the K⁺-evoked [³H]noradrenaline release from superfused rat brain cortical and hippocampal synaptosomes. In cortical presynaptic terminals the 1,4-dihydropyridine agonist Bay K 8644 enhanced the K⁺ (15 m M )-evoked [³H]noradrenaline release. This effect was reversed by the 1,4-dihydropyridine antagonists nimodipine and nitrendipine. The L-type VSCC ligands had no effect on hippocampal synaptosomes. In contrast, the N-type VSCC blocker ω-conotoxin markedly reduced the K⁺-evoked [³H]noradrenaline release in nerve terminals from both regions. Inhibition was greater in hippocampal synaptosomes. When applied together the inhibitory actions of nimodipine and ω-conotoxin were approximately additive. These findings indicate that both L- and N-type VSCCs participate in noradrenaline release in rat brain cortex and suggest that noradrenergic terminals in the two regions examined may have distinct populations of VSCCs: L type in cortex and N type in hippocampus.     

Neurochemical characterization of the striatum and the nucleus accumbens in L-type Ca(v)1.3 channels knockout mice

L-type Ca(v)1.3 channels control the autonomous pacemaking of the substantia nigra (SN) dopamine (DA) neurons, which maintains the sustained release of DA in the striatum, its target structure. The persistent engagement of L-type channels during pacemaking might lead to increased vulnerability to environmental stressors or degenerative processes, providing a mechanism for the development of Parkinson's disease (PD). Interestingly, L-type channels are not necessary for pacemaking, opening the possible use of calcium channel antagonists as neuroprotective agents for PD without disturbing normal DA function. In this study we aimed to evaluate the consequences of Ca(v)1.3 channels deletion at the neurochemical level. For this purpose, tissue concentrations of DA and their respective metabolites were measured using high performance liquid chromatography (HPLC) in the striatum and the nucleus accumbens (NAcc) of mice lacking the gene for the Ca(v)1.3 channel subunit (CACNA1D) and compared to those in wild-type mice. Striatal DA level did not differ between the two groups. In contrast, the level of serotonin, glutamate, GABA, and taurine were increased by more than 50% in the striatum of Ca(v)1.3 null mice. Neurotransmitters levels in the NAcc did not differ between the different groups. In conclusion, our results neurochemically corroborate the robustness of the nigrostriatal DA neurons in the absence of Ca(v)1.3 channels, but suggest that complete deletion of this channel affected a variety of other transmitter systems.     

Decreased calcium accumulation in isolated nerve endings during hibernation in ground squirrels

Resting and depolarization-induced⁴⁵CaCl₂ accumulation was compared for synaptosomes isolated from hibernating and nonhibernating ground squirrels. Channel subtype antagonists were used to identify the active voltage-sensitive calcium channel subtypes in these preparations. There was significantly less⁴⁵Ca²⁺ accumulation in synaptosomes isolated from hibernating as compared to cold-adapted nonhibernating ground squirrels in both basal (p<0.005) and depolarizing (p<0.03) media over a 30 sec to 5 min incubation period. The elevation in⁴⁵Ca²⁺ accumulation triggered by K⁺ depolarization was blocked by 50 μM CdCl₂, 1 μM ω-conotoxin MVIIC or 1 μM ω-agatoxin IVA. Inhibition was not observed with 1 μM nifedipine or with 1 μM ω-conotoxin GVIA. These results suggest that hibernation is associated with reduced presynaptic⁴⁵Ca²⁺ conductance via voltage-sensitive channels with a pharmacological sensitivity that is different from the established L-, N-, and P-types in other systems but share features of the recently described Q-type calcium channel. This decrease may reflect a cellular adaptation that helps confer tolerance to the near total cerebral ischemia associated with hibernation.     

Effect of nicotine on extracellular levels of neurotransmitters assessed by microdialysis in various brain regions: Role of glutamic acid

We studied the effect of local administration of nicotine on the release of monoamines in striatum, substantia nigra, cerebellum, hippocampus, cortex (frontal, cingulate), and pontine nucleus and on the release of glutamic acid in striatum of rats in vivo, using microdialysis for nicotine administration and for measuring extracellular amine and glutamic acid levels. Following nicotine administration the extracellular concentration of dopamine, increased in all regions except cerebellum; serotonin increased in cingulate and frontal cortex; and norepinephrine increased in substantia nigra, cingulate cortex, and pontine nucleus. Cotinine, the major nicotine metabolite, had no effect at similar concentrations. The cholinergic antagonists mecamylamine and atropine, the dopaminergic antagonists haloperidol and sulpiride, and the excitatory amino acid antagonist kynurenic acid all inhibited the nicotine-induced increase of extracellular dopamine in the striatum. The fact that kynurenic acid almost completely prevented the effects of nicotine, and nicotine at this concentration produced a 6-fold increase of glutamic acid release, suggests that the effect of nicotine is mainly mediated via glutamic acid release.     

Striatal GDNF administration increases tyrosine hydroxylase phosphorylation in the rat striatum and substantia nigra

Glial cell line-derived neurotrophic factor (GDNF) improves motor dysfunction associated with aging in rats and non-human primates, in animal models of Parkinson's disease, and may improve motoric function in patients with advanced Parkinson's disease. These improvements are associated with increased dopamine function in the nigrostriatal system, but the molecular events associated with this increase are unknown. In these studies, 100 micro g of GDNF was injected into the striatum of normal aged (24-month-old) male Fischer 344 rats. The protein levels and phosphorylation of TH, ERK1/2, and related proteins were determined by blot-immunolabeling of striatum and substantia nigra harvested 30 days after injection. In GDNF-treated rats, TH phosphorylation at Ser31 increased approximately 40% in striatum and approximately 250% in the substantia nigra. In the substantia nigra, there was a significant increase in ERK1 phosphorylation. In striatum, there was a significant increase in ERK2 phosphorylation. Microdialysis studies in striatum showed that both amphetamine- and potassium-evoked dopamine release in GDNF recipients were significantly increased. These data show that GDNF-induced increases in dopamine function are associated with a sustained increase in TH phosphorylation at Ser31, which is greatest in the substantia nigra and maintained for at least one month following a single striatal administration of GDNF. These findings, taken from the nigrostriatal system of normal aged rats, may help explain the long lasting effects of GDNF on dopamine function and prior studies supporting that a major effect of GDNF involves its effects on dopamine storage and somatodendritic release of dopamine in the substantia nigra.     

Biotransformation of l-DOPA to Dopamine in the Substantia Nigra of Freely Moving Rats: Effect of Dopamine Receptor Agonists and Antagonists

Abstract: We investigated the effects of continuous intranigral perfusion of dopamine D1 and D2 receptor agonists and antagonists on the biotransformation of locally applied l -DOPA to dopamine in the substantia nigra of freely moving rats by means of in vivo microdialysis. The "dual-probe" mode was used to monitor simultaneously changes in extracellular dopamine levels in the substantia nigra and the ipsilateral striatum. Intranigral perfusion of 10 µ M l -DOPA for 20 min induced a significant 180-fold increase in extracellular nigral dopamine level. No effect of the intranigral l -DOPA administration was observed on dopamine levels in the ipsilateral striatum, suggesting a tight control of extracellular dopamine in the striatum after enhanced nigral dopamine levels. Continuous nigral infusion with the D1 receptor agonist CY 208243 (10 µ M ) and with the D2 receptor agonist quinpirole at 10 µ M (a nonselective concentration) attenuated the l -DOPA-induced increase in dopamine in the substantia nigra by 85 and 75%, respectively. However, perfusion of the substantia nigra with a lower concentration of quinpirole (1 µ M ) and the D1 antagonist SCH 23390 (10 µ M ) did not affect the nigral l -DOPA biotransformation. The D2 antagonist (−)-sulpiride (10 µ M ) also attenuated the l -DOPA-induced dopamine release in the substantia nigra to ∼10% of that of the control experiments. We confirm that there is an important biotransformation of l -DOPA to dopamine in the substantia nigra. The high concentrations of dopamine formed after l -DOPA administration may be the cause of dyskinesias or further oxidative stress in Parkinson's disease. Simultaneous administration of D1 receptor agonists with l -DOPA attenuates the biotransformation of l -DOPA to dopamine in the substantia nigra. The observed effects could occur via changes in nigral GABA release that in turn influence the firing rate of the nigral dopaminergic neurons.     

Antigens Associated with N- and L-Type Calcium Channels in Lambert-Eaton Myasthenic Syndrome

Abstract: In Lambert-Eaton myasthenic syndrome neurotransmitter release is reduced by an autoimmune response directed against the calcium channel complex of the nerve terminal. Autoantibodies were detected by immunoprecipitation assays using solubilized receptors labeled with ligands selective for N-type (¹²⁵I-ω conotoxin GVIA) and L-type ([³H]PN200-110) calcium channels. Sera with a high antibody titer (>3 n M ) against rat brain N-type channels contained autoantibodies that immunoprecipitated neuronal and muscle L-type channels. These IgG fractions stained a 55-kDa protein in immunoblots of purified skeletal muscle dihydropyridine receptor, suggesting that they contain autoantibodies against the β subunit of the calcium channel. A distinct antibody population in the same fractions reacted with a nerve terminal 65-kDa protein that is unrelated to the β subunit and displays properties similar to those of synaptotagmin.     

N-type calcium channels are involved in the dopamine releasing effect of nicotine

Mouse striatum was incubated with [³H]dopamine ([³H]DA) and superfused with and the tritium efflux induced by nicotine, electrical stimulation, or simultaneous nicotine and electrical stimulation was measured, to characterize the role of different Ca²⁺ channels in the transmitter release. Nicotine stimulation and electrical stimulation exerted additive effects on tritium efflux. Separation of the released radioactivity on alumina columns indicated that nicotine or electrical stimulation increases the release of [³H]DA and that the outflow of³H-labeled metabolites was similar with the two different stimulation procedures. Removal of Ca²⁺ from the superfusate resulted in a marked reduction in the tritium release evoked by nicotine, whereas the electrical stimulation-evoked tritium release was completely dependent on external Ca²⁺. The L-and N-type calcium channel blockers omega-conotoxin GVIA and Cd²⁺ inhibited the tritium release from the striatum evoked by either nicotine or electrical stimulation, whereas the L-type and T-type channel blockers diltiazem and Ni²⁺ did not alter release of [³H]DA. We conclude that N-type voltage-sensitive Ca²⁺ channels participate in striatal dopamine release, and we speculate that nicotinic receptor-operated ion channels permeable to cations such as Ca²⁺ and N-type voltage-sensitive calcium channels may simultaneously open up, and they additively increase free intracellular Ca²⁺ concentration.     

3-Methoxytyramine Formation Following Monoamine Oxidase Inhibition Is a Poor Index of Dendritic Dopamine Release in the Substantia Nigra

Abstract: This study was undertaken, using microdialysis, to compare the extracellular concentration of 3-methoxytyramine and dopamine in dialysate from the striatum and substantia nigra, after pargyline (75 mg/kg), after pargyline plus amphetamine (3 mg/kg), and after pargyline plus reserpine (5 mg/kg) administration. Treatment with pargyline alone increased the extracellular dopamine concentration by 70% in the striatum and by 140% in the substantia nigra and induced in both regions a time-dependent accumulation of 3-methoxytyramine. The addition of d-amphetamine to pargyline increased the extracellular dopamine concentration, compared with pargyline-treated controls, to the same extent in both the substantia nigra (maximally by 360%) and the striatum (maximally by 400%), but the concomitant increase of 3-methoxytyramine accumulation in the dialysate was relatively smaller in the substantia nigra compared with the striatum. Reserpine treatment decreased the extracellular dopamine concentration in both regions below the detection level (<10% of basal value). When pargyline was added to reserpine, the striatal extracellular dopamine concentration increased to 50% of pargyline-treated controls and the striatal 3-methoxytyramine accumulation was less than in pargyline-treated controls. However, in the substantia nigra, the addition of pargyline to reserpine resulted in dopamine concentrations as high as after pargyline only and the 3-methoxytyramine accumulation was not changed compared with pargyline-treated controls. In summary, our results indicate that dopamine in the substantia nigra is released from reserpine-sensitive storage sites and that pargyline-induced 3-methoxytyramine accumulation is a poor indicator of the local dopamine release. The latter observation may be explained by the fact that the dopamine-metabolizing enzyme, catechol-O-methyltransferase, is located inter alia in the dopamine-containing cell bodies/dendrites in the substantia nigra, in contrast to the situation in the terminals in the striatum where catechol-O-methyltransferase is located only in nondopaminergic cells.     

Different Effects of Opiate Withdrawal on Dopamine Turnover,Uptake, and Release in the Striatum and Nucleus Accumbens

The purpose of these experiments was to further characterize changes in dopaminergic function that follow withdrawal from chronic opiate treatment. Withdrawal after treatment to a maximum dose of 120 mg/kg of morphine did not alter dopamine concentrations in the substantia nigra, ventral tegmental area, striatum, or nucleus accumbens; but did decrease concentrations of DOPAC and the ratio of DOPAC to dopamine in the lateral striatum and nucleus accumbens. Uptake of tritiated dopamine was diminished for withdrawn slices obtained from the striatum with no effect observed for tissue from the nucleus accumbens. Deficits of in vitro release of tritiated dopamine also occurred following withdrawal, with the nucleus accumbens being sensitive to dependence produced by a lower dose of morphine. In conclusion, opiate withdrawal produces a complex pattern of effects on dopaminergic function that is specific for the striatum and nucleus accumbens.     

Limited regulation of somatodendritic dopamine release by voltage-sensitive Ca channels contrasted with strong regulation of axonal dopamine release

The mechanism underlying somatodendritic release of dopamine (DA) appears to differ from that of axon-terminal release. Specifically, somatodendritic DA release in the substantia nigra pars compacta (SNc) persists in low extracellular Ca2+ concentrations that are insufficient to support axonal release in striatum, suggesting that limited Ca2+ entry is necessary to trigger somatodendritic release. Here, we compared the role of voltage-dependent Ca2+ channels in mediating DA release in striatum versus SNc using specific blockers of N-, P/Q-, T-, R- and L-type Ca2+ channels individually and in combination. Release of DA evoked by a single stimulus pulse in the dorsal striatum and SNc of guinea-pig brain slices was monitored in real time using carbon-fiber microelectrodes with fast-scan cyclic voltammetry. Single-pulse evoked DA release was shown to be independent of regulation by concurrently released glutamate or GABA acting at ionotropic receptors in both regions. Under these conditions, striatal DA release was completely prevented by an N-type channel blocker, omega-conotoxin GVIA (100 nm), and was decreased by 75% by the P/Q-type channel blocker omega-agatoxin IVA (200 nm). Blockade of T-type channels with Ni2+ (100 microm) or R-type channels with SNX-482 (100 nm) decreased axonal release in striatum by 25%, whereas inhibition of L-type channels with nifedipine (20 microm) had no effect. By contrast, none of these Ca2+-channel blockers altered the amplitude of somatodendritic DA release in the SNc. Even a cocktail of all blockers tested did not alter release-signal amplitude in the SNc, although the duration of the release response was curtailed. The limited involvement of voltage-dependent Ca2+ channels in somatodendritic DA release provides further evidence that minimal Ca2+ entry is required to trigger the release process, compared with that required for axon-terminal release.     

Novel Pharmacological Sensitivity of the Presynaptic Calcium Channels Controlling Acetylcholine Release in Skate Electric Organ

Abstract: The presynaptic terminals of skate ( Raja montagui ) electric organ were tested for their sensitivity to calcium channel antagonists. Acetylcholine (ACh) release and the elevation of intraterminal Ca²⁺ concentrations triggered by K⁺ depolarisation were studied. ACh release was measured as ³H efflux from slices of organ prelabelled with [³H]choline. Depolarisation caused a marked, Ca²⁺-dependent increase in ³H efflux that was completely blocked by 100 µ M Cd²⁺ and by 300 n M ω-conotoxin-MVIIC (MVIIC). Inhibition by MVIIC was concentration dependent (IC₅₀ of ∼20 n M ) and reversible. No inhibition was seen with nifedipine (5 µ M ) or the two other peptide antagonists studied: ω-conotoxin-GVIA (GVIA) at 5 µ M and ω-agatoxin-IVA (Aga-IVA) at 1 µ M . In a "nerve plate" preparation (a presynaptic plexus of nerve fibres, Schwann cells, and nerve terminals) changes in intraterminal Ca²⁺ concentrations were measured by microfluorimetry using fluo-3. An increase in fluorescence, indicating a rise in the free [Ca²⁺], rapidly followed K⁺ depolarisation, and this change was restricted to the nerve terminals. This response was insensitive to nifedipine (5 µ M ), GVIA (5 µ M ), and Aga-IVA (300 n M ) but almost completely abolished by MVIIC (1 µ M ). MVIIC inhibition was concentration dependent and partially reversible. These results show that the nerve terminals in skate electric organ have calcium channels with a pharmacological sensitivity that is markedly different from the established L, N, and P types in other systems but shares some, but not all, of the features of the recently described Q type.     

Responsiveness of nigral neurons to the stimulation of striatal dopaminergic receptors in the rat

The microinfusion of low doses of apomorphine into the striatum of anesthetized rats depressed the electrical activity of the neurons of the substantia nigra pars compacta while the infusion of bromocriptine had an excitatory or inhibitory effect. These data suggest that:1) the action of the two dopamine agonists on the striato-nigral pathway is different; 2) the striatum might contain dopaminergic receptors located on cells projecting to the substantia nigra with different roles in the feedback regulation of the latter; 3) the inhibitory action of systemically injected apomorphine is not simply due to a stimulation of dopamine “autoreceptors” but also to an action mediated by fibers descending from the striatum to the substantia nigra.