George K. Aghajanian

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).

The formation of synaptic junctions in developing rat brain: A quantitative electron microscopic study

Abstract The formation of synaptic junctions in developing rat brain has been studied with the electron microscope, utilizing a selective staining method. In this procedure, glutaraldehyde-fixed tissue is not exposed to osmium tetroxide, but instead is blockstained with phosphotungstic acid in solution of ethanol. A selective staining of the paramembranous components of synaptic junctions results from this treatment, thus facilitating a quantification of the numerical density of synaptic junctions over large areas of sections. It was found that a sharp increase in the number of synaptic junctions occurs in rat cortex (molecular layer) during the 3rd and 4th postnatal weeks. The relationship between this increase in the numerical density of synaptic junctions and other events during development, such as the maturation of the EEG, is discussed. In addition, it is proposed that synaptic junctions evolve through several stages and a tentative morphogenetic scheme is presented.

Catecholamine receptors on locus coeruleus neurons: Pharmacological characterization

Abstract There is evidence that the noradrenergic neurons of the locus coeruleus (LC) possess α-adrenoreceptors in the vicinity of their cell bodies. To further characterize this receptor, we studied the responses of LC neurons to a series of catecholamine agonist and antagonist drugs using the techniques of single-unit recording and microiontophoresis. The spontaneous firing of LC neurons was inhibited by microiontophoretic application of norepinephrine, epinephrine, the α-adrenoreceptor agonist clonidine and the β-agonist isoproterenol. These inhibitions were blocked by the α-adrenergic antagonist piperoxane but not by the β-antagonist sotalol. In addition these cells were strongly inhibited by dopamine or α-methylnorepinephrine, but only weakly inhibited by phenylephrine or the dopamine agonist apomorphine. The dopamine antagonist trifluoperazine was ineffective in blocking the inhibitions of LC neurons by both dopamine and norepinephrine. The rank order of potencies of agonist drugs in inhibiting LC neurons was clonidine > > α-methylnorepinephrine ⩾ epinephrine = norepinephrine > > phenylephrine. In this respect the LC α-receptor is similar to ‘presynaptic’ or ‘α2’ receptors or peripheral sympathetic nerves and differs from the classical postsynaptic α-receptor. The noradrenergic neurons of the LC thus appear to possess catecholamine receptors on or near their cell bodies which have pharmacological characteristics of ‘presynaptic’ α-adrenergic receptors. The LC receptors are distinct from central dopamine receptors and from norepinephrine receptors areas of the brain receiving their noradrenergic iput fom the LC.

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”.

Glutamate N-methyl-D-aspartate Receptor Antagonists Rapidly Reverse Behavioral and Synaptic Deficits Caused by Chronic Stress Exposure

BACKGROUND Despite widely reported clinical and preclinical studies of rapid antidepressant actions of glutamate N-methyl-D-aspartate (NMDA) receptor antagonists, there has been very little work examining the effects of these drugs in stress models of depression that require chronic administration of antidepressants or the molecular mechanisms that could account for the rapid responses. METHODS We used a rat 21-day chronic unpredictable stress (CUS) model to test the rapid actions of NMDA receptor antagonists on depressant-like behavior, neurochemistry, and spine density and synaptic function of prefrontal cortex neurons. RESULTS The results demonstrate that acute treatment with the noncompetitive NMDA channel blocker ketamine or the selective NMDA receptor 2B antagonist Ro 25-6981 rapidly ameliorates CUS-induced anhedonic and anxiogenic behaviors. We also found that CUS exposure decreases the expression levels of synaptic proteins and spine number and the frequency/amplitude of synaptic currents (excitatory postsynaptic currents) in layer V pyramidal neurons in the prefrontal cortex and that these deficits are rapidly reversed by ketamine. Blockade of the mammalian target of rapamycin protein synthesis cascade abolishes both the behavioral and biochemical effects of ketamine. CONCLUSIONS The results indicate that the structural and functional deficits resulting from long-term stress exposure, which could contribute to the pathophysiology of depression, are rapidly reversed by NMDA receptor antagonists in a mammalian target of rapamycin dependent manner.

Antidromic identification of dopaminergic and other output neurons of the rat substantia nigra

In the present study dopamine (DA)-containing and other output neurons of the substantia nigra (SN) wer identified by antidromic stimulation from postulated target nuclei, the caudate-putamen, the thalamus, the cortex and the pontine reticular formation. To guide electrode placements, the topography of the nigrostriatal projection system was determined by retrograde tracing methods. Spontaneously active cells present in the SN were then classified in two groups according to the shape of their action potentials and their firing rate. Type I cells were located mainly in the pars compacta and could be antidromically-activated (AD-activated) from various locations along the course of the nigrostriatal pathway (caudate-putamen, globus pallidus, MFB) but not from other brain areas (ventromedial thalamus, motor cortex, pontine reticular formation). These neurons had a slow bursting pattern of firing, a very slow conduction velocity (0.58 m/sec), and a wide action potential. Their firing rate was dramatically reduced following the intravenous administration of apomorphine (ID 50: 9.3 microgram/kg), or the iontophoretic application of DA and GABA. Type II cells were located predominantly in the pars reticulata; most of them could be AD-activated from the ventromedial thalamus and the MFB but not from the motor cortex. A few of these cells could be AD-activated from the pontine reticular formation and the thalamus. A minority of type II cells, located in or near the pars compacta could be AD-activated from the caudate-putamen. In addition, their conduction velocuty was much higher (2.8 m/sec) and their firing rate far in excess of that exhibited by type I neurons. These neurons were inhibited by the iontophoretic application of GABA but not of DA. The microinjection of 6-hydroxydopamine (a neurotoxin relatively specific against catecholamine-containing neurons) in the vicinity of the MFB blocked selectively the propagation of antidromic spikes in type I but not type II cells. It is concluded that type I cells are the DA neurons of the nigrostriatal pathway. Type II cells are mainly oupput neurons that project to the ventromedial thalamus, the pons and the forebrain. This telencephalic projection most likely constitutes a second, non-DA, fast-conducting nigrostriatal pathway.

Synaptic Dysfunction in Depression: Potential Therapeutic Targets

Basic and clinical studies demonstrate that depression is associated with reduced size of brain regions that regulate mood and cognition, including the prefrontal cortex and the hippocampus, and decreased neuronal synapses in these areas. Antidepressants can block or reverse these neuronal deficits, although typical antidepressants have limited efficacy and delayed response times of weeks to months. A notable recent discovery shows that ketamine, a N-methyl-d-aspartate receptor antagonist, produces rapid (within hours) antidepressant responses in patients who are resistant to typical antidepressants. Basic studies show that ketamine rapidly induces synaptogenesis and reverses the synaptic deficits caused by chronic stress. These findings highlight the central importance of homeostatic control of mood circuit connections and form the basis of a synaptogenic hypothesis of depression and treatment response.

Electrophysiological and pharmacological characterization of serotonergic dorsal raphe neurons recorded extracellularly and intracellularly in rat brain slices

Extracellular and intracellular recordings were made from dorsal raphe (DR) neurons in frontal rat brain slices maintained in vitro. A population of neurons was found which displayed electrophysiological and pharmacological characteristics of serotonin-containing DR neurons recorded in vivo. Recorded extracellularly, these neurons displayed biphasic or triphasic action potentials of 1.5-3.0 ms duration, and discharged with a slow and steady rhythm. Recorded intracellularly these neurons displayed action potentials of about 1.8 ms duration, which were followed by large (10-20 mV) after hyperpolarizations which normally lasted 200-800 ms. These presumed serotonergic DR neurons were inhibited by LSD and serotonin. They were excited by norepinephrine, or the alpha-agonist phenylephrine, and these activations could be reduced or blocked by alpha-adrenoreceptor antagonists including the selective alpha 1-antagonist, prazosin. The major difference between the in vitro recordings and previous in vivo recordings from anesthetized animals was a reduction in the number of spontaneously firing DR neurons. This was probably due, at least in part, to a disfacilitation of serotonergic DR neurons in the slice caused by the functional removal of a tonic noradrenergic input.

Serotonergic facilitation of facial motoneuron excitation.

The effect of serotonin (5-HT) on motoneurons located in the facial nucleus of the rat was investigated in the present study. Microiontophoretic application of 10--200 nA pulses of 5-HT lasting from 1 to 10 min failed to excite facial motoneurons. However, small amounts of 5-HT facilitated the subthreshold and threshold excitatory effects of iontophoretically applied glutamate on these cells. Typically, the current of glutamate required to produce an activation of facial motoneurons was reduced by at least 50% in the presence of 5-HT. In addition, 5-HT markedly shifted to the left the cumulative dose-response curve of glutamate-induced excitation of motoneurons. The 5-HT releasing agent p-chloroamphetamine (PCA) facilitated the excitatory effects of glutamate on montoneurons in control animals, but not in those pretreated with the 5-HT also facilitated the subthreshold and threshold excitation of motoneurons produced by stimulation of the motor cortex and the red nucleus. The facilitating effect of 5-HT was blocked by methysergide. Norepinephrine also facilitated facial motoneuron excitation but this effect was not blocked by methysergide. It is concluded that 5-HT in the facial nucleus functions in a manner that is not analagous to direct excitation, but rather acts as a gain setter to enhance the effects of excitatory afferent inputs.

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.