Zona Compacta Neurons Research Articles

Differential actions of AMP kinase on ATP-sensitive K+ currents in ventral tegmental area and substantia nigra zona compacta neurons.

ATP-sensitive K+ (K-ATP) channels play significant roles in regulating the excitability of dopamine neurons in the substantia nigra zona compacta (SNC). We showed previously that K-ATP channel function is up-regulated by AMP-activated protein kinase (AMPK). This study extended these studies to the neurons adjacent to the SNC in the ventral tegmental area (VTA). Using patch pipettes to record whole-cell currents in slices of rat midbrain, we found that the AMPK activator A769662 increased the amplitude of currents evoked by the K-ATP channel opener diazoxide in presumed dopamine-containing VTA neurons. However, current evoked by diazoxide with A769662 was significantly smaller in VTA neurons compared to SNC neurons. Moreover, a significantly lower proportion of VTA neurons responded to diazoxide with outward current. However, A769662 was able to increase the incidence of diazoxide-responsive neurons in the VTA. In contrast, A769662 did not potentiate diazoxide-evoked currents in presumed non-dopamine VTA neurons. These results show that AMPK activation augments K-ATP currents in presumed dopamine neurons in the VTA and SNC, although diazoxide-evoked currents remain less robust in the VTA. We conclude that K-ATP channels may play important physiological roles in VTA and SNC dopamine neurons.

Rotenone reduces Mg 2+-dependent block of NMDA currents in substantia nigra dopamine neurons

Rotenone is a pesticide that has been successfully used to produce a rodent model of Parkinson's disease. We reported previously that rotenone potently augmented N-methyl- d-aspartate (NMDA)-evoked currents in rat dopamine neurons via a tyrosine kinase-dependent mechanism. In this study, we investigated the effect of rotenone on the current–voltage relationship of NMDA-induced currents in substantia nigra zona compacta neurons recorded with whole-cell patch pipettes in slices of rat brain. In a physiologic concentration of extracellular Mg 2+ (1.2 mM), a 30 min perfusion with rotenone (100 nM) produced a marked increase in current evoked by bath application of NMDA (30 μM), especially when measured at relatively hyperpolarized currents. In the presence of rotenone, NMDA currents lost the characteristic region of negative-slope conductance that is normally produced by voltage-dependent block by Mg 2+. The voltage-dependent effect of rotenone on NMDA currents was mimicked by a low extracellular concentration of Mg 2+ (0.2 mM) and was antagonized by a high level of Mg 2+ (6.0 mM). Moreover, we report that the tyrosine kinase inhibitor genistein blocked the ability of rotenone to augment NMDA receptor currents. These results suggest that rotenone potentiates NMDA currents by a tyrosine kinase-dependent process that attenuates voltage-dependent Mg 2+ block of NMDA-gated channels. These results support the hypothesis that an excitotoxic mechanism might participate in rotenone-induced toxicity of midbrain dopamine neurons.

Baclofen inhibition of the hyperpolarization-activated cation current, Ih, in rat substantia nigra zona compacta neurons may be secondary to potassium current activation.

1. The properties of the hyperpolarization-activated cation current (Ih), and its modulation by gamma-aminobuturic acid-B (GABAB) receptor activation and protein kinase A, were investigated using whole cell voltage clamp of substantia nigra zona compacta principal neurons in rat midbrain slices in vitro. 2. At 30 degrees C, Ih activated between -75 and -155 mV, with a V1/2 of -115 mV. At 35 degrees C, the activation curve shifted positive by 10 mV. Ih had an estimated reversal potential of -27 mV. Ion substitution experiments showed that the current was carried by Na+ and K+. 3. Application of the GABAB receptor agonist baclofen (30 microM) induced an outward potassium current (GIRK), increased neuronal membrane conductance and inhibited Ih. The inhibition of Ih was voltage independent. Baclofen induced an 11-mV positive shift in the reversal potential of Ih. 4. Extracellular barium (300 microM) markedly reduced the baclofen-evoked outward current and associated increase in membrane conductance due to GIRK activation. There was also very little inhibition of Ih by baclofen in the presence of barium. When cesium was the major intracellular cation, both the increase in membrane conductance due to GIRK activation and the inhibition of Ih evoked by baclofen were reduced by a similar extent. 5. Neither forskolin (10 microM) nor the protein kinase A inhibitor, H89 (10 microM), had any effect on Ih or its inhibition by baclofen. 6. These data suggest that the inhibition of Ih by baclofen is secondary to the activation of GIRK, i.e., due directly to alteration of membrane conductance, rather than a distinct effect, and is not mediated by inhibition of adenylyl cyclase.

Organization and physiology of the substantia nigra.

A comprehensive review of the literature on the anatomy, electrophysiology and pharmacology of the substantia nigra is presented. A diagram is developed taking into account the interneuronal interactions of neurotransmitters and receptors that control firing rates and neurotransmitter releases. The central features of the diagram are a positive dopaminergic feedforward process and a positive feedback mechanism mediated by extrasynaptic substance P diffusing from striatal terminals to dopaminergic dendrites of the zona compacta neurons. This loop can enhance the transmission of information from the striatum through the pars reticulata output neurons. The loop is controlled at the level of zona compacta neurons by a negative feedback supported by the dendritic release of dopamine and boosted by pedunculopontine activation mediated by muscarinic receptors. The output of the loop is controlled by two negative feedforward processes, both involving GABAergic striatonigral afferents. Application of the model to pharmacological studies of diverse behaviors including seizures, turning, and conditioned behaviors reveals unforseen relationships and may offer insights into, and directions for, further analysis of the mechanisms and functions involved.

The selective NK3 receptor agonist senktide excites a subpopulation of dopamine-sensitive neurones in the rat substantia nigra pars compacta in vitro.

Intra- and extracellular recordings were made from substantia nigra zona compacta neurones from an in vitro rat brain slice preparation. A subpopulation of dopamine-sensitive neurones were encountered which were potently excited by bath application of the NK3 receptor agonist, senktide. On these senktide-sensitive neurones, NK1 and NK2 receptor agonists were inactive. The excitatory action of senktide supports a role for tachykinins as putative neurotransmitters in the basal ganglia.

Nomifensine but not amantadine increases dopamine-induced responses on rat substantia nigra zona compacta neurons

Responses of substantia nigra zona compacta neurons to nomifensine and amantadine were studied with intracellular recording techniques (current and voltage clamp) in in vitro slice preparation of rat mesencephalon. The application of nomifensine (1–10 μM) slightly hyperpolarized the cells and inhibited action potential discharge that occurs spontaneously. In voltage-clamp experiments (−50, −60 mV, holding potential) an outward current was observed. The membrane responses to exogenously-applied dopamine were potentiated by the concomitant superfusion of nomifensine. The effects of nomifensine were antagonized by (−)-sulpiride (1 μM), a D 2 receptor antagonist. By contrast, the superfusion of amantadine (1–30 μM) on substantia nigra zona compacta cells was ineffective on firing rate, membrane potential or on sensitivity to exogenous dopamine. In the presence of high doses (300 μM to 1 mM) of amantadine a depolarization and an increase in firing activity was observed. While our results provide electrophysiological evidence for an inhibition of the dopamine uptake system by nomifensine, they do not support a dopaminergic mechanism for the actions of amantadine in the substantia nigra zona compacta.

The electrophysiological properties of substantia nigra pars compacta neurones recorded from 6-hydroxydopamine lesioned guinea-pigs in vitro

Intracellular recordings were made from substantia nigra pars compacta neurones in vitro from animals with partial unilateral 6-hydroxydopamine lesions of the nigrostriatal tract. Lesions were assessed and grouped according to the severity of the striatal dopamine depletion. No differences were seen between neurones from control and lesioned side nigrae as regards their membrane properties, firing rates, burst activity or percentage of quiescent neurones in any of the lesioned categories. It is concluded that following partial lesioning, the remaining substantia nigra zona compacta neurones in vitro, are functioning normally.

Dopamine uptake inhibition potentiates the effects of l-DOPA on rat substantia nigra zona compacta neurons

Intracellular electrophysiological recordings (current and voltage-clamp mode) were used to asses whether the actions of l-DOPA on rat substantia nigra zona compacta neurons maintained in vitro are potentiated by drugs that block the dopamine uptake system (cocaine and nomifensine). Bath application of l-DOPA (100 μM) hyperpolarized the dopaminergic cells and inhibited the spontaneous occurring action potential discharge. Under voltage-clamp experiments, l-DOPA produced an outward current that was related to the hyperpolarization of the membrane. In the presence of 10 μM cocaine and 10 μM nomifensine, the cellular responses to l-DOPA application were augmented and prolonged. In contrast to this, amphetamine (a dopamine releasing drug), did not augment the membrane response to l-DOPA. Cocaine (10 μM), nomifensine (10 μM) and amphetamine (3 μM), slightly hyperpolarized the membrane potential and inhibited the firing activity of the cells. It is concluded that the dopamine uptake system have an important role in regulating the cellular responses (membrane hyperpolarization or outward current) to l-DOPA application in the substantia nigra zona compacta.

Responses of rat substantia nigra compacta neurones to L‐DOPA

1. The electrophysiological actions of L-DOPA were studied on substantia nigra dopamine-containing neurones by use of intracellular recordings, current and voltage clamp, in vitro. 2. L-DOPA (3-300 microM), applied by superfusion, decreased the spontaneous firing activity of the neurones, hyperpolarized the membrane potential and produced an outward current. 3. These effects of L-DOPA were graded and were antagonized by sulpiride, a D2-receptor antagonist. 4. The L-DOPA activated membrane hyperpolarization persisted in slices exposed to tetrodotoxin while it was markedly reduced or abolished in solutions with no calcium plus cobalt (2 mM). 5. In slices treated with carbidopa, an inhibitor of the aromatic amino acid decarboxylase enzyme, the actions of L-DOPA were greatly reduced. 6. We propose that L-DOPA inhibits the firing of substantia nigra zona compacta neurones by being converted to dopamine which is then released by these cells.

Actions of cocaine on rat dopaminergic neurones in vitro

1. Intracellular recordings were made from neurones in vitro in the rat substantia nigra zona compacta and ventral tegmental area; the neurones had the characteristic properties of dopamine-containing cells, and fired action potentials spontaneously. 2. Cocaine (1-10 microM) inhibited spontaneous firing, hyperpolarized the membrane and (in neurones voltage-clamped at -60 mV) caused an outward membrane current; the minimally effective concentration was 1 microM. These effects were blocked by sulpiride (30 nM-1 microM). 3. Dopamine (3-100 microM) also inhibited firing, hyperpolarized and caused an outward current. These effects of dopamine were potentiated about five fold by cocaine (10 microM). 4. It is concluded that cocaine (1-10 microM) inhibits the firing and hyperpolarizes substantia nigra zona compacta neurones in vitro by blocking the uptake of dopamine which the cells continuously release.

Effects of glycine on neurons in the rat substantia nigra zona compacta: In vitro electrophysiological study

The effects of bath-applied glycine to substantia nigra zona compacta neurons of rat were investigated by intracellular recording techniques in vitro. Superfusion of glycine (1 mM) in the medium hyperpolarized 53% of the neurons recorded with KCl electrodes, whereas 32% of the cells were depolarized. The remaining 15% of neurons was hyperpolarized and then depolarized by the amino acid. In spite of these membrane changes, the action potential firing was depressed. Both hyperpolarization and depolarization were correlated to an outward and an inward current, respectively, when recording in single-electrode voltage-clamp mode. In response to bath application of glycine, the neurons showed a concentration-dependent conductance increase. Micromolar concentrations of glycine (100-300 microM) in the superfusion medium produced a membrane hyperpolarization (outward current) in most of the tested cells, whereas millimolar concentration of amino acid could cause depolarization (inward current) in the same neurons. When the recording electrodes contained K acetate, only hyperpolarizations (outward current) were produced. The potential and current changes and the increase in membrane conductance produced by glycine showed little desensitization when neurons were recorded with K acetate electrodes. The mean reversal potential for the membrane hyperpolarization was -80 mV with intracellular electrodes containing KCl and -84 mV with electrodes containing K acetate. The mean null potential for the depolarizing effect was -46 mV. The reversal potential for the glycinergic responses was shifted to less negative values upon lowering the extracellular concentration of chloride or increasing the extracellular concentration of potassium. Strychnine (1-10 microM) reversibly antagonized both the conductance increase and the membrane changes produced by glycine. Bath application of bicuculline (100 microM) and picrotoxin (200 microM) did not affect glycine responses, while depressing the actions of GABA and muscimol. It is concluded the glycine exerts an inhibition on substantia nigra zona compacta neurons by acting on strychnine-sensitive receptors. The membrane effects produced by glycine result from the activation of a chloride current. In addition, the simultaneous involvement of potassium ions may contribute to the overall effects of glycine.

The mechanism of amphetamine-induced inhibition of rat substantia nigra compacta neurones investigated with intracellular recording in vitro.

1. The effects of (+)-amphetamine (1-300 microM) applied by superfusion, on substantia nigra zona compacta neurones were studied in slices of the rat mesencephalon with intracellular recording techniques (current and voltage clamp mode). 2. Amphetamine inhibited the action potential discharge, hyperpolarized the cells and increased the membrane conductance. Under voltage clamp it produced an outward current. 3. The amphetamine outward current reversed at potentials negative to -100 mV (2.5 mM extracellular potassium) and was reduced by barium, (1 mM) suggesting an increased potassium conductance. 4. In slices treated with tetrodotoxin and calcium-free solution and added cobalt, amphetamine was still effective. 5. (-)-Sulpiride, (1 microM) a D2 antagonist, blocked the effects of both amphetamine and dopamine. 6. In slices treated for more than 3 h with 30 microM alpha-methyl-DL-tyrosine, an agent known to block the synthesis of dopamine, the action of amphetamine was attenuated, while dopamine was still effective. 7. Responses to applied dopamine were potentiated by cocaine (10 microM) but unaffected by amphetamine (1-3 microM). 8. It is suggested that the inhibition of substantia nigra compacta neurones by amphetamine is mainly due to the release of endogenous dopamine.

Two cell types in rat substantia nigra zona compacta distinguished by membrane properties and the actions of dopamine and opioids

Intracellular recordings were made from 475 rat substantia nigra zona compacta neurons in vitro. The region from which recordings were made was rich in catecholamine fluorescence. Two groups of neuron, termed principal neurons (95% of the total) and secondary neurons (5% of the total) were clearly distinguishable according to one or more of the following 4 electrophysiological properties. Secondary neurons (23 cells) (1) fired spontaneous action potentials at frequencies greater than 10 Hz, or were quiescent (30%); (2) had action potentials less than 1 msec in duration; (3) did not show time-dependent inward rectification with step hyperpolarization; and (4) had slope conductances of about 4 nS (between -75 and -90 mV). In contrast, principal neurons (1) fired spontaneous action potentials in the range 1-8 Hz, or were quiescent (33%); (2) had action potentials greater than 1 msec in duration; (3) showed pronounced time-dependent inward rectification; and (4) had steady-state membrane slope conductances of around 22 nS (between -75 and -90 mV). Secondary cells were not affected by dopamine but were hyperpolarized by baclofen, GABA, and the mu opioid receptor agonist Tyr-D-Ala-Gly-MePhe-Gly-ol (DAGO). On the other hand, dopamine and baclofen inhibited firing and/or hyperpolarized all principal cells tested, but mu or delta opioid receptor agonists had no effect. The properties of these 2 cell types broadly correspond with those described by electrophysiological studies in vivo, in which case the majority, or principal, cells are believed to be dopaminergic.(ABSTRACT TRUNCATED AT 250 WORDS)

Electrophysiological evidence for the dendritic localization of a calcium conductance in guinea-pig substantia nigra neurones in vitro.

Within the substantia nigra, anatomical, neurochemical and pharmacological findings strongly suggest that transmitter and protein are secreted from the dendrites of nigrostriatal neurones. This phenomenon may underlie a non classical modulatory cellular mechanism. Two conductances are generated in nigrostriatal neurones independent of somatic action potentials, that might mediate this modulation. However, these conductances have never been directly nor precisely located specifically within the dendrites. The aim of this study was to record the membrane properties of substantia nigra zona compacta neurones in response to selective sectioning of the population of long 'apical' dendrites i.e. the removal of the zona reticulata. Intracellular recordings from substantia nigra zona compacta neurones were made from mesencephalic slices of the guinea-pig brain maintained in vitro. In cells without the apical dendrites, the membrane potential, input resistance and mean firing frequency was not significantly different from the control neurones. However, removal of the substantia nigra zona reticulata virtually abolished one conductance in particular. This conductance, seen in control neurones, is a long lasting slow depolarization which is resistant to tetrodotoxin blockade of sodium channels: rather, it is mediated by the entry of calcium ions and is optimally deinactivated at a hyperpolarised membrane potential. Hence, this study strongly suggests that this conductance is generated exclusively in the 'apical' dendrites. It has been postulated that this long lasting calcium conductance is central to the modulation of nigrostriatal neuronal excitability. Thus, the 'apical' dendrites could play a specific and active role in the functioning of nigrostriatal neurones.

A possible pacemaker mechanism in pars compacta neurons of the guinea-pig substantia nigra revealed by various ion channel blocking agents

The membrane properties of pars compacta neurons in the in vitro guinea-pig substantia nigra have been studied in the presence of sodium, calcium and potassium channel blockers. The following properties, which have already been described for dopamine-containing substantia nigra zona compacta neurons were observed: high and low threshold calcium spikes: a calcium-activated potassium-mediated transient; inward rectification. Inward rectification was sensitive to caesium ions. An additional property was seen reminiscent of an “A” current, although resistant to 4-aminopyridine. It is suggested that this outward transient is in fact a calcium activated potassium conductance. Under certain conditions calcium-mediated rhythmic depolarizations were observed. It is suggested that at least two of the properties seen (outward rectification and low threshold calcium spike) could interact to provide the basis for a pacemaker mechanism in pars compacta neurons.

Potassium ions play a role in the glycine-induced inhibition of rat substantia nigra zona compacta neurones

Glycine directly inhibits the firing, it hyperpolarizes and/or depolarizes the dopaminergic neurones and increases the membrane conductance. In voltage clamp experiments (near resting potential) either outward and/or inward currents were produced. These actions were present in tetrodotoxin and in 0-calcium-cobalt-treated slices and were antagonized by strychnine. The hyperpolarization depended on the extracellular concentration of potassium and was prevented by cesium diffusion into the cell. Thus potassium ions participate in the glycine mediated inhibition of the firing of substantia nigra zona compacta cells.

On the potassium conductance increase activated by GABAB and dopamine D2 receptors in rat substantia nigra neurones.

1. Intracellular recordings were made from 193 substantia nigra zona compacta neurones in slices of rat mesencephalon. All cells were hyperpolarized by baclofen; this was accompanied by a fall in input resistance. Cells voltage clamped at -60 mV showed an outward current associated with a conductance increase in response to baclofen. The baclofen effects were concentration dependent (effective range 0.3-30 microM); the concentration producing half the maximal effect was 1.5 microM. (-)-Baclofen was 300-700 times more potent than (+)-baclofen. 2. The potential change or membrane current caused by baclofen reversed polarity at -108.8 +/- 1.1 mV (n = 10) when the potassium ion concentration was 2.5 mM, -96.0 +/- 2.8 mV (n = 3) in 4.5 mM-potassium and -76.6 +/- 1.7 mV (n = 5) in 10.5 mM-potassium. The relationship between reversal potential and potassium concentration conformed to the Nernst equation. 3. Dopamine was also applied to 119 of these neurones; all exhibited either a hyperpolarization or an outward current. 4. Baclofen and dopamine outward currents were reduced reversibly by barium (100-300 microM) and tetraethylammonium (10 mM). Superfusion for 5-10 min with solutions presumed to block calcium currents reduced, but did not abolish, responses to baclofen. The effect of baclofen persisted in tetrodotoxin (1 microM). 5. Superfusion of gamma-aminobutyric acid (GABA, 0.3-3 mM) caused either membrane depolarization or hyperpolarization, accompanied by a fall in input resistance. The depolarization was mimicked by muscimol (10 microM) and blocked by bicuculline methiodide (10-100 microM); the hyperpolarization was resistant to bicuculline. Nipecotic acid (500 microM) enhanced the effect of GABA, but was without effect upon the actions of muscimol and baclofen. 6. The effect of dopamine was enhanced by cocaine (10 microM) and antagonized by (-)-sulpiride (0.1-1 microM), whereas the actions of baclofen were unaffected by cocaine or (-)-sulpiride. The maximum outward current produced by dopamine was approximately half that produced by baclofen. 7. Outward currents produced by dopamine were reversibly occluded by maximal outward currents caused by baclofen. 8. Baclofen and dopamine hyperpolarizations were unaffected by intracerebroventricular injection of animals with pertussis toxin. 9. Cells impaled with electrodes containing guanosine 5'-O-(3-thiotriphosphate) (1 mM) were hyperpolarized by both baclofen and dopamine, but the membrane potential did not fully return to its original level when agonist application was discontinued. 10. It is concluded that activation of both dopamine D2 and GABAB receptors may increase the same potassium conductance.(ABSTRACT TRUNCATED AT 400 WORDS)

Cocaine modulation of central monoaminergic neurotransmission

Extracellular microelectrode studies were conducted to test the effects of cocaine HCl on the activity of spontaneously firing single serotonergic dorsal raphe (DRN), noradrenergic locus coeruleus (LC) and dopaminergic ventral tegmental (VTA) and zona compacta (ZC) neurons, and cerebellar Purkinje neurons (PC) in urethane anesthetized rats in vivo. Cocaine (0.0625-4 mg/kg) predominantly inhibited all of the central monoaminergic neurons and predominantly activated cerebellar Purkinje neurons. Cocaine (1 mg/kg, IV) failed to potentiate the inhibitory effects of LC stimulation on PC neurons. The temporal effects of intravenous cocaine on arterial pressure (i.e., pressor response) were not directly correlated with the effects on neurons. Cocaine did not decrease the amplitude or slope of neuron action potentials, and the effects of cocaine on firing rate were not shared by similar doses of procaine. Reserpine pretreatment (10 mg/kg, IP) attenuated the effects of cocaine (1 mg/kg, IV) on DRN, LC, and PC neurons. Specific adrenoceptor antagonists antagonized the inhibitory effects of cocaine on LC (piperoxane, yohimbine) and VTA (haloperidol) neurons. These results suggest that the central effects of cocaine on presynaptic monoaminergic neurons may in part be mediated by augmented monoamine neurotransmission at autoreceptors and that the effects of cocaine on postsynaptic target cells (PC) may be more complex, requiring the analysis of both pre- and postsynaptic elements.

Hyperpolarizing action of baclofen on neurons in the rat substantia nigra slice

GABA, muscimol and l-baclofen inhibited zona compacta neurons in the rat substantia nigra slice. d-baclofen was inactive, l-baclofen produced a potent hyperpolarization of the membrane potential. The actions of muscimol but not GABA and l-baclofen were reversed by bicuculline. The actions of muscimol and baclofen are likely to be mediated by interactions with two different receptors.

Electrophysiological properties of mouse dopamine neurons: In vivo and in vitro studies

The present experiments were conducted to determine the electrophysiological and pharmacological properties of substantia nigra neurons in the mouse. These cells were studied using extracellular single unit recording and microiontophoretic techniques in both chloral hydrate anesthetized mice and in vitro mouse slices. In the in vivo preparation the substantia nigra zona compacta neurons had long duration action potentials (> 4 ms), fired from 1 to 7 impulses/s, and the cells discharged with either a decremental burst pattern or with a regular pattern. The dopamine agonists apomorphine and d-amphetamine, given systemically, decreased the firing rate of these neurons and the dopamine receptor blocker, haloperidol, reversed these effects. The zona compacta neurons were inhibited by the micro-iontophoretic application of dopamine and γ-aminobutyric acid, and systemic haloperidol selectively attenuated the effects of dopamine. In vitro recordings from substantia nigra zona compacta and zona reticulata neurons were generally similar to those found in vivo, both in terms of the electrophysiological and pharmacological properties. However, the zona compacta cells fired faster in vitro than in vivo, and the firing pattern in vitro tended to be pacemaker-like, especially when recordings were made in an incubation medium which blocks synaptic transmission (e.g. low Ca 2+/high Mg 2+). Our data indicate that: (a) in vivo mouse zona compacta neurons exhibit the same electrophysiological and pharmacological properties as rat dopamine-containing neurons; (b) in vitro mouse dopaminergic neurons fire with pacemaker regularity when in a low Ca 2+/high Mg 2+ environment; and (c) in vitro studies offer an approach to examine the basic properties of dopaminergic neurons exclusive of feedback pathways and other afferent inputs.