Zhi Gen Jiang

Membrane properties and synaptic responses of rat striatal neurones in vitro.

1. A tissue slice containing a section of striatum was cut obliquely from rat brain so as to preserve adjacent cortex and pallidum. Intracellular recordings were made from 368 neurones, using either conventional or tight‐seal configurations. 2. Two types of neurone were distinguished electrophysiologically. Principal cells (96%) had very negative resting potentials (‐89 mV) and a low input resistance at the resting membrane potential (39 M omega): membrane conductance (10 nS at ‐65 mV) increased within tens of milliseconds after the onset of hyperpolarization (99 nS at ‐120 mV). Secondary cells (4%) had less negative resting potentials (‐60 mV) and a higher input resistance (117 m omega at the resting potential): hyperpolarization caused an inward current to develop over hundreds of milliseconds that had the properties of H‐current. 3. Most principal cells were activated antidromically by electrical stimulation of the globus pallidus or internal capsule. Intracellular labelling with biocytin showed that principal cells had a medium sized soma (10‐18 microns), extensive dendritic trees densely studded with spines and, in some cases, a main axon which extended towards the globus pallidus. 4. Electrical stimulation of the corpus callosum or external capsule evoked a depolarizing postsynaptic potential. This synaptic potential was reversibly blocked by a combination of 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX, 10 microM) and DL‐2‐amino‐5‐phosphonovaleric acid (APV, 30 microM), but was unaffected by bicuculline (30 microM) and picrotoxin (100 microM). The underlying synaptic current had a fast component (time to peak about 4 ms), the amplitude of which was linearly related to membrane potential and which was blocked by CNQX; in CNQX the synaptic current had a slower component (time to peak about 10 ms) which showed voltage dependence typical of N‐methyl‐D‐aspartate (NMDA) receptors. Both currents reversed at ‐5 mV. 5. Focal electrical stimulation within the striatum (100‐300 microns from the site of intracellular recording) evoked a synaptic potential that was partially blocked (45‐95%) by CNQX and APV: the remaining synaptic potential was blocked by bicuculline (30 microM). The bicuculline‐sensitive synaptic current reversed at the chloride equilibrium potential. 6. The findings confirm that the majority of neostriatal neurones (principal cells, medium spiny neurones) project to the pallidum and receive synaptic inputs from cerebral cortex mediated by an excitatory amino acid acting through NMDA and non‐NMDA receptors. These cells also receive synaptic inputs from intrinsic striatal neurones mediated by GABA.(ABSTRACT TRUNCATED AT 400 WORDS)

Actions of 5-hydroxytryptamine on ventral tegmental area neurons of the rat in vitro

Intracellular recordings were made with conventional microelectrodes and with whole-cell patch-clamp electrodes from neurons of the rat ventral tegmental area and substantia nigra zona compacta in vitro. Neurons were distinguished as principal cells and secondary cells; it is known from previous work that most principal cells contain dopamine whereas secondary cells do not. 5-Hydroxytryptamine (5-HT; 3-100 microM) depolarized (or evoked an inward current at -60 mV) 46% of 153 principal cells; a small proportion (11%) of cells were hyperpolarized (or showed outward current at -60 mV). Secondary cells were equally likely to be depolarized (or inward current at -60 mV, 30% of 80 cells) or hyperpolarized (or outward current at -60 mV, 28%). approximately 40% of each type of cell were unaffected by 5-HT. Depolarizing responses of 5-HT were mimicked by (+/-)-1-(2,5-dimethoxy-4-iodophenyl)- 2-aminopropane (DOI) and blocked by ketanserin. Hyperpolarizing responses were mimicked by dipropyl-5-carboxamidotryptamine and reversed polarity at the K+ equilibrium potential. Inhibitory postsynaptic potentials (or currents) mediated at GABAA receptors occurred spontaneously in some principal cells; they were reversibly blocked by tetrodotoxin and bicuculline. 5-HT either increased or decreased the frequency of these synaptic potentials but did not change their mean amplitude or decay time.(ABSTRACT TRUNCATED AT 250 WORDS)

Membrane properties and synaptic inputs of suprachiasmatic nucleus neurons in rat brain slices.

1. Whole‐cell recordings were made from 390 neurons of the suprachiasmatic nucleus (SCN) in horizontal brain slices during different portions of the circadian day. The locomotor activity of the rats was measured prior to the preparation of brain slices to insure that each rat was entrained to a 12 h‐12 h light‐dark cycle. 2. The mean input conductance was 42% higher (1.58 nS) in neurons recorded near the subjective dawn than those (1.11 nS) recorded near the subjective dusk. The current required to hold the neurons at ‐60 mV also showed a circadian variation with a peak in the middle of the subjective day and a nadir in the middle of the subjective night. Analysis of the variations in the input conductance and the holding current at ‐60 mV suggested that at least two ion conductances are involved in the pacemaking of the circadian rhythms. 3. Voltage‐clamped SCN neurons often had both outward and inward spontaneous postsynaptic currents. The outward currents were blocked by bicuculline but not by strychnine, and were identified as IPSCs mediated by GABAA receptors. The inward currents were blocked by 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX) and were identified as EPSCs mediated by glutamate. Most spontaneous synaptic currents were miniature currents but action potential‐dependent large events were seen more often in IPSCs than in EPSCs. 4. Stimulation of the optic nerve or chiasm usually evoked a monosynaptic EPSC which was mediated by both NMDA and non‐NMDA receptors. In 13% of cells, optic nerve stimulation evoked an outward current or an inward current followed by an outward current; all the evoked currents were blocked by 4‐aminophosphonovaleric acid (APV) and CNQX whereas the outward current only was blocked by bicuculline, suggesting involvement of an inhibitory interneuron. 5. SCN neurons sum the excitatory inputs from both optic nerves; on average each SCN cell receives innervation from at least 4.8 retinohypothalamic tract (RHT) axons. 6. Focal stimulation in the vicinity of the recorded neuron revealed that nearly all SCN neurons receive local or extranuclear GABAergic inputs operating via GABAA receptors. The EPSCs activated by such stimulation were not significantly different in amplitude and pharmacological properties from those induced by RHT stimulation. 7. One hundred and one neurons were labelled with neurobiotin during whole‐cell recording. Based on the dendritic structures, four types of SCN neurons (monopolar, radial, simple bipolar and curly bipolar) were identified. The curly bipolar cells had a higher membrane conductance, holding current and hyperpolarization‐activated current (Ih) amplitude than the other neuronal types. Radial neurons did not respond to optic nerve stimulation, which activated EPSCs in the other cell types.

Melatonin activates an outward current and inhibits Ih in rat suprachiasmatic nucleus neurons

Whole-cell voltage-clamp recordings were made from suprachiasmatic nucleus (SCN) neurons maintained in horizontal brain slices. The majority of neurons exhibited spontaneous and evoked excitatory and inhibitory synaptic currents (EPSC and IPSC), mediated by glutamate and GABA respectively. Melatonin had no effect on either the spontaneous or evoked EPSC or IPSC. Application of melatonin (0.1-30 microM) during circadian time (CT) 9-12 activated an outward current at -60 mV and increased the membrane conductance in a concentration-dependent manner. The current was augmented by depolarization, reduced by hyperpolarization and, in some cells, reversed its polarity near the potassium equilibrium potential. Some neurons also responded to melatonin during other times of the circadian day (CT 3-9 or CT 12-15). Hyperpolarizing steps, in a portion of cells, activated an inward cation current which resembled the Ih described in other neurons. Melatonin (10 microM) inhibited activation of the Ih. These data indicate that melatonin may inhibit SCN neurons by activating a potassium current and inhibiting the Ih.

PRESYNAPTIC INHIBITION BY BACLOFEN OF RETINOHYPOTHALAMIC EXCITATORY SYNAPTIC TRANSMISSION IN RAT SUPRACHIASMATIC NUCLEUS

Optic nerve stimulation evoked monosynaptic excitatory postsynaptic currents in suprachiasmatic nucleus neurons maintained in vitro. These currents were completely blocked by a combination of glutamate receptor antagonists, 6-cyano-7-nitroquinoxaline-2,3-dione and 4-aminophosphonovaleric acid. Stimulation of the ipsilateral or contralateral suprachiasmatic nucleus produced a biphasic response consisting of an excitatory postsynaptic current followed by an bicuculline-sensitive inhibitory postsynaptic current. Most suprachiasmatic nucleus neurons had spontaneous inhibitory and excitatory synaptic currents produced by action potential-independent and, less frequently, action potential-dependent release of GABA and glutamate. Baclofen reversibly reduced the amplitude of excitatory postsynaptic currents evoked by optic nerve stimulation and the effect was antagonized by 2-hydroxysaclofen. In addition, baclofen reduced the frequency but not the amplitude of the spontaneous miniature excitatory postsynaptic currents. In a subset of suprachiasmatic nucleus neurons, baclofen induced an outward current, probably by increasing a potassium conductance. Baclofen had no effect on either evoked or spontaneous inhibitory postsynaptic currents or on currents activated by pulse application of glutamate. These data indicate that activation of GABAB receptors can inhibit suprachiasmatic nucleus neurons by two mechanisms. The first is to inhibit the release of glutamate from terminals of the retinohypothalamic tract. The second is the postsynaptic activation of a potassium conductance in a portion of these neurons.

Pre- and postsynaptic actions of serotonin on rat suprachiasmatic nucleus neurons

Serotoninergic transmission is implicated in the photic and non-photic regulation of circadian rhythms. 5-HT (1-100 microM), carboxamidotryptamine (5-CT 0.1-10 microM) and (+)-8-hydroxy-dipropylaminotetraline (8-OH-DPAT, 1-30 microM) dose-dependently activated an outward current (5-100 pA) in 30% of neurons voltage-clamped at -60 mV in the suprachiasmatic nucleus (SCN) in vitro slice. EC(50) values were 7.0 microM for 5-HT and 0.2 microM for 5-CT. Serotonin-induced outward current was associated with an increase in input conductance, and the current was blocked by Ba(2+) (1 mM). The amplitude of the current was enhanced by depolarization, reduced by hyperpolarization, and reversed its polarity during a hyperpolarization beyond the potassium equilibrium potential. Mean amplitudes of the 5-HT outward current changed with time of the subjective circadian day. The value near CT2 (23.8 pA) was about 4 times greater than that around CT14 (6.7 pA). Cells that responded with an outward current showed four types of morphology: monopolar, simple bipolar, curly bipolar and radial shaped; they were localized in all parts of the SCN. The EPSC evoked by retino-hypothalamic-tract (RHT) stimulation was inhibited 26% but the inward current induced by exogenously applied glutamate or NMDA was not affected by serotonin agonists. Focal stimulation-induced and spontaneous IPSC but not the exogenous GABA-induced outward current were inhibited by 5-HT agonists in a subpopulation of cells. In conclusion, 5-HT regulates SCN neurons by both pre- and post-synaptic inhibitory mechanisms; the latter may play a key role in modulating SCN circadian rhythm by activation of 5-HT receptors and opening of a potassium channel.

Synaptic Transmission and Signal Transduction in the Suprachiasmatic Nucleus

Circadian rhythms are daily patterns of physiological processes which persist in the absence of environmental time cues, indicating the existence of an endogenous biological clock. In humans, the circadian clock plays a important role in sleep-wake cycles, jet lag, and the performance ability of shift workers. The circadian clock of mammals is located in the suprachiasmatic nucleus (SCN), which is composed of nuclei located bilaterally of the third ventricle and dorsal to the optic chiasm, each consisting of approximately 8,000 neurons (Van den Pol 1980).