Benjamin S. Bunney

Inhibition of both noradrenergic and serotonergic neurons in brain by the α-adrenergic agonist clonidine

By means of single unit recording techniques it was found that a small systemically administered (intravenous) dose of the alpha-adrenergic agonist clonidine inhibited the spontaneous firing of brain norepinephrine (NE)-containing neurons in the locus coeruleus. In addition, the NE neurons were consistently inhibited by the direct (microiontophoretic) application of minute amounts of NE or clonidine. Intravenous clonidine also inhibited the firing of the great majority of (5-HT) neurons in the midbrain dorsal raphe nucleus. However, this action does not appearto be a direct one since clonidine (and NE) had relatively weak or variable effects when applied microiontophoretically to raphe neurons. The clonidine-induced depression of raphe firing may be secondary to an impairment in adrenergic transmission since (1) the depression could be reversed by the NE-releasing agents D- and L-amphetamine, (2) high doses of clonidine itself (which have been reported to have postsynaptic alpha-agonistic activity) reversed the depression produced by a low dose of clonidine and (3) prior destruction of NE neurons by 6-hydroxydopamine (7-12 days) rendered raphe neurons insensitive to the depressant effect of i.v. clonidine. Dopaminergic (substantia nigra, zona compacta) neurons did not respond to either low or high doses of clonidine. These results are consistent with previous data showing that clonidine decreases NE and 5-HT but not dopamine turnover. We conclude that systemically administered clonidine inhibits the firing of brain NE neurons by acting directly upon adrenergic receptors located on or near the soma of these neurons but that the concomitant inhibition of 5-HT neurons is an indirect effect (possibly secondary to an impairment in noracrenergic transmission).

Intracellular and extracellular electrophysiology of nigral dopaminergic neurons—1. Identification and characterization

Intracellular recordings were obtained from directly identified rat nigral dopamine cells in vivo. This identification was based on an increase in glyoxylic acid-induced catecholamine fluorescence in the impaled dopamine neurons. One of three compounds was injected intracellularly into each cell to produce the heightened fluorescence: (1) L-DOPA, to increase the intracellular dopamine content by precursor loading; (2) tetrahydrobiopterin, a cofactor for tyrosine hydroxylase, to increase intracellular dopamine concentration through activation of the rate-limiting enzyme for dopamine synthesis and (3) colchicine, to arrest intraneuronal transport and thus allow the build-up of dopamine synthesizing enzymes and dopamine in the soma. In addition, dopamine cells were antidromically activated from the caudate nucleus and collision with a directly elicited action potential was demonstrated. Identified dopamine neurons were shown to possess an input resistance of 31.2 +/- 7.4 M omega (means +/- SD) and a time constant of 12.1 +/- 3.2 ms. The action potentials were of long duration (2.75 +/- 0.5 ms) with a marked break between the initial segment and the somatodendritic spike components. The initial segment was the only component commonly elicited during antidromic activation. Spontaneously occurring action potentials were usually preceded by a slow, pacemaker-like depolarization. Burst firing by summation of depolarizing afterpotentials was observed to occur spontaneously, but could not be triggered by short depolarizing current pulses. Intravenously administered apomorphine demonstrated the same inhibitory effect on cell firing that was previously reported to occur when recording extracellularly from identified dopaminergic neurons. The determination of the electrophysiological characteristics of a population of cells directly identified as containing a specific neurotransmitter (in this case, dopamine) may allow one to construct better models of a systems functioning. Thus, the high input resistance and long time constant of dopamine-containing cells, combined with their burst/pause firing mode, may be important functionally with respect to a possible modulatory effect of dopamine in postsynaptic target areas.

Typical and atypical neuroleptics: differential effects of chronic administration on the activity of A9 and A10 midbrain dopaminergic neurons

Extracellular single unit recording techniques were used to study the effects of both acute and repeated oral neuroleptic administration on the in vivo activity of rat A9 and A10 dopaminergic (DA) neurons. All antipsychotic drugs examined acutely (haloperidol, l-sulpiride, chlorpromazine, and clozapine) increased the number of spontaneously firing DA neurons in both A9 and A10 compared to controls. Repeated (21 day) treatment with haloperidol, l-sulpiride, and chlorpromazine (antipsychotic drugs which can cause extrapyramidal side effects) markedly reduced the number of active DA cells below control levels in both regions. The “silent” DA neurons were in an apparent state of tonic depolarization inactivation since they could be induced to discharge by the microiontophoretic application of the inhibitory neurotransmitter gamma-aminobutyric acid, but not the excitatory amino acid glutamate. The depolarization inactivation observed may be specific for antipsychotic drugs since a non-neuroleptic phenothiazine (promethazine), the inactive isomer of sulpiride (d-sulpiride), and a tricyclic antidepressant (desmethylimipramine) neither increased DA activity when given acutely nor induced depolarization inactivation when administered repeatedly. In contrast to the other drugs tested, repeated treatment with clozapine (an effective antipsychotic drug which does not produce extrapyramidal side effects) resulted in the depolarization inactivation of A10 neurons but not A9 cells. These data suggest that neuroleptics which can induce extrapyramidal side effects produce depolarization inactivation of both A9 and A10 neurons whereas antipsychotic drugs which lack this property inactivate only A10 neurons. It is suggested that the time-dependent development of A9 DA neuron inactivation induced by repeated neuroleptic treatment may provide a mechanism for understanding the delayed onset of extrapyramidal side effects often observed with these drugs.

Dopamine “Autoreceptors”: Pharmacological characterization by microiontophoretic single cell recording studies

SummaryThe effects on the firing of single dopamine (DA) neurons in the substantia nigra (and adjacent ventral tegmental area) of a representative group of catecholamine agonists and antagonists were studied in rats using single cell recording and microiontophoretic techniques. Microiontophoretic application of DA or the DA agonist apomorphine depressed the firing of these cells; the DA antagonist trifluoperazine blocked this effect. However, the α-agonist clonidine had no depressant effect and the β-agonist isoproteronol had only a weak depressant action on DA neurons. Furthermore, the α-antagonist piperoxane and the β-antagonist sotolol were completely ineffective in blocking the depressant effects of DA. These results show that DA-sensitive receptors on the soma of DA neurons are pharmacologically distinct from α or β adrenoreceptors. Because of their location and selective responsiveness to DA agonists, the catecholamine receptors on the soma of DA neurons appear best classified as DA “autoreceptors”.

Acute and chronic haloperidol treatment: Comparison of effects on nigral dopaminergic cell activity

Abstract Antipsychotic drugs produce most of their clinical effects, both therapeutic and adversive, in a time-dependent manner which, depending upon the effect, can take days to years to develop. Using extracellular single unit recording and microiontophoretic techniques, we investigated the effect of chronic haloperidol (CHAL) treatment (0.5 mg/kg/day s.c. × 22 d) on nigral dopaminergic (DA) neuronal activity. These effects were compare to those obtained in control animals, animals acutely treated with haloperidol (AHAL), and animals which had been treated for 21 days but not tested until a week after haloperidol had been discontinued (CHAL+l). CHAL treatment resulted in an almost total absence of spontaneously firing nigral DA cells. “Silent” DA cells became active when GABA or DA was applied microiontophoretically but they were unresponsive to glutamic acid. I.V. apomorphine also caused the DA cells to fire. Destruction of nigro-striatal feedback pathways by injection of kainic acid into the caudate nucleus prior to CHAL treatment prevented the disappearance of dopamine cell activity on the lesioned side. In AHAL animals a significantly greater number of spontaneously firing DA cells were found than in controls. In control animals inhibited DA cells could be activated by microiontophoretic glutamic acid or i.v. haloperidol but not by GABA. These results suggest that CHAL treatment causes an increase in the activity of DA cells to the point that the great majority go into apparent tonic depolarization block. This effect appears to be mediated via striato-nigral feedback pathways. AHAL treatment appears to activate DA cells that are normally inactive as well as accelerate the firing rate of spontaneously firing DA neurons. The possible relevance of these findings to the time-dependent neurological side effects induced by haloperidol is discussed.

Noradrenergic neurons: morphine inhibition of spontaneous activity.

Abstract The effect of morphine sulfate on the spontaneous firing rate of norepinephrine-containing neurons in the locus coeruleus was studied in rats. Morphine sulfate was found to selectively inhibit neuronal activity in the locus coeruleus but had no effect on the firing rate of serotonergic neurons in the dorsal raphe nucleus. Naloxone, a morphine antagonist, blocked and reversed the morphine-induced inhibition of locus coeruleus cells. Chlorpromazine, in contrast to its anti-amphetamine effects, did not antagonize the depression of spontaneous activity of the NE cells by morphine. Naloxone was ineffective in blocking or reversing d-amphetamine inhibition of noradrenergic neuron activity. A noxious stimulus (toe pressure) transiently increased the firing rate of neurons in the locus coeruleus. This response was markedly reduced by morphine sulfate. These findings suggest that morphine may exert at least part of this analgesic action through decreasing locus coeruleus impulse flow. However, the mechanism by which morphine decreases noradrenergic neuronal activity remains to be elucidated.

Acute Effects of Typical and Atypical Antipsychotic Drugs on the Release of Dopamine from Prefrontal Cortex, Nucleus Accumbens, and Striatum of the Rat: An In Vivo Microdialysis Study

Abstract: In vivo microdialysis has been used to study the acute effects of antipsychotic drugs on the extracellular level of dopamine from the nucleus accumbens, striatum, and prefrontal cortex of the rat. (–)‐Sulpiride (20, 50, and 100 mg/kg i.v.) and haloperidol (0.1 and 0.5 mg/kg i.v.) enhanced the outflow of dopamine in the striatum and nucleus accumbens. In the medial prefrontal cortex, (–)‐sulpiride at all doses tested did not significantly affect the extracellular level of dopamine. The effect of haloperidol was also attenuated in the medial prefrontal cortex; 0.1 mg/kg did not increase the outflow of dopamine and the effect of 0.5 mg/kg haloperidol was of shorter duration in the prefrontal cortex than that observed in striatum and nucleus accumbens. The atypical antipsychotic drug clozapine (5 and 10 mg/kg) increased the extracellular concentration of dopamine in all three regions. In contrast to the effects of sulpiride and haloperidol, that of clozapine in the medial prefrontal cortex was profound. These data suggest that different classes of antipsychotic drugs may have distinct effects on the release of dopamine from the nigrostriatal, mesolimbic, and mesocortical terminals.

The precise localization of nigral afferents in the rat as determined by a retrograde tracing technique.

Afferent innervation of the rat substantia nigra (SN) was studied by the retrograde horseradish peroxidase (HRP) method. High concentrations of HRP were deposited in discrete subregions of the SN by means of a microiontophoretic delivery system. Using this technique it was possible to demonstrate that the caudatonigral projection system is arranged topographically; All portions of the caudate-putamen except for a central medial core were found to contain HRP positive cells, indicative of retrograde transport. In the positive areas a much larger percentage of cells (30-50%) were found to participate in this projection than has previously been reported. Only medium size cells (12-20 mum) were found to contain the HRP reaction product. Other areas found to heavily innervate the SN were the globus pallidus, central nucleus of the amygdala and dorsal raphe nucleus. Areas containing fewer reactive cells but which also appear to innervate the SN included the prefrontal cortex and lateral habenula. These results emphasize the importance of striatonigral projections which recent studies have suggested contain a GABAergic link.

Peptide-monoamine coexistence: studies of the actions of cholecystokinin-like peptide on the electrical activity of midbrain dopamine neurons.

Abstract Recent studies have shown that a cholecystokinin-like peptide coexists in a subpopulation of midbrain dopamine-containing neurons. Using extracellular single unit recording techniques we have found that this peptide increases the activity of some, but not all, dopaminergic neurons (identified on the basis of their electrophysiological features). The responsive neurons were found exclusively in areas which were subsequently shown, using immunocytochemical techniques, to contain both cholecystokinin and the enzyme marker for dopaminergic neurons, tyrosine hydroxylase. When administered intravenously, cholecystokinin increased the firing rate of dopaminergic neurons lying in cholecystokinin-rich areas of the substantia nigra. Cells in cholecystokinin-rich ventral tegmental areas showed more variable responses to comparable doses of the peptide. Iontophoretically-applied cholecystokinin consistently activated dopaminergic cells in the substantia nigra and ventral tegmental area, increasing their firing rate and their bursting activity. Cholecystokinin increased the firing rate of some of those neurons to the extent that they were driven into apparent depolarization inactivation. Furthermore, iontophoresis of cholecystokinin resulted in an activation of a population of normally quiescent dopaminergic cells. These results are discussed in light of a possible functional role of cholecystokinin-like peptides in the brain dopaminergic systems. It is suggested that cells in the dopamine-rich areas of the mesencephalon can be characterized both on the basis of their content of peptide and/or catecholamine and of their responsiveness to cholecystokinin-like peptides.

Evidence for the absence of impulse-regulating somatodendritic and synthesis-modulating nerve terminal autoreceptors on subpopulations of mesocortical dopamine neurons

Electrophysiological and biochemical techniques were used to study midbrain dopamine systems. In the electrophysiological studies, projection areas of individual dopaminergic cells were identified by antidromic activation. Dopamine cells which innervate the piriform cortex and those that innervate the caudate nucleus demonstrated their usual dose-dependent inhibitory response to both the intravenous administration of the direct-acting dopamine agonist apomorphine and the microiontophoretic application of dopamine. In contrast, the firing rate of dopamine neurons which project to the prefrontal cortex and of those terminating in the cingulate cortex was not altered by either the intravenous administration of low to moderate doses of apomorphine or microiontophoretically applied dopamine. The mean basal discharge rate and degree of burst firing was also different between these subgroups of midbrain dopaminergic neurons. Mesoprefrontal and mesocingulate dopamine neurons had mean firing rates of 9.3 and 5.9 spikes/s respectively, and showed intense burst activity. Mesopiriform and nigrostriatal dopamine cells had discharge rates of 4.3 and 3.1 spikes/s and displayed only moderate bursting. The dopaminergic nature of those mesocortical neurons insensitive to apomorphine and dopamine was confirmed using combined intracellular recording and catecholamine histofluorescence techniques. Thus, after the intracellular injection of colchicine and subsequent processing for glyoxylic acid-induced histofluorescence, the injected cells could be identified by their brighter fluorescences compared to the surrounding, normally fluorescing, non-injected dopamine neurons. Using biochemical techniques, subgroups of midbrain dopaminergic systems were again found to differ. The administration of gamma-butyrolactone increased dopamine levels in all areas sampled (prefrontal, cingulate and piriform cortices as well as the caudate nucleus). However, although this effect was readily reversed in both the piriform cortex and caudate nucleus by pretreatment with apomorphine, this treatment had no effect on the increased dopamine levels observed in the prefrontal and cingulate cortices. In addition, the decline in dopamine levels after synthesis inhibition with alpha-methyltyrosine was significantly faster in the prefrontal and cingulate cortices relative to the caudate nucleus. The piriform cortex showed an intermediate decline which was not significantly different from that observed in any of the other regions.(ABSTRACT TRUNCATED AT 400 WORDS)