Reduction In Input Resistance Research Articles

Pre- and postsynaptic gaba b receptors modulate rapid neurotransmission from suprachiasmatic nucleus to parvocellular hypothalamic paraventricular nucleus neurons

The suprachiasmatic nucleus (SCN), the dominant circadian pacemaker in mammalian brain, sends axonal projections to the hypothalamic paraventricular nucleus (PVN), a composite of magno- and parvocellular neurons. This neural network likely offers SCN output neurons a means to entrain diurnal rhythmicity in various autonomic and neuroendocrine functions. Earlier investigations using patch-clamp recordings in slice preparations have suggested differential innervation by SCN efferents to magnocellular versus parvocellular PVN cells. In magnocellular PVN, cells respond to focal electrical stimulation in SCN with a GABA A receptor-mediated postsynaptic inhibition whose magnitude can be modulated by presynaptic GABA B receptors. By contrast, SCN-evoked responses in parvocellular PVN neurons typically involve both GABA A- and glutamate-receptor-mediated components. In the present patch-clamp study, 69/85 periventricular parvocellular PVN cells displayed SCN-evoked inhibitory and/or excitatory postsynaptic currents (IPSCs; EPSCs). In the presence of selective receptor antagonists, we sought evidence for their modulation by GABA acting at pre- and/or postsynaptic GABA B receptors. Cells responded to bath-applied baclofen (5–10 μM) with a tetrodotoxin-resistant membrane hyperpolarization associated with a reduction in input resistance and/or outward current, due to increase in a potassium conductance, blockable with 2-hydroxysaclofen (300 μM). At 1 μM where baclofen had no significant postsynaptic effect, evidence of activation of presynaptic GABA B receptors included reduction in SCN-evoked IPSCs and EPSCs with no change in their kinetics, and paired-pulse depression that was sensitive to both baclofen and saclofen. Baclofen also induced significant reductions in frequency but not amplitudes of miniature IPSCs and EPSCs. These observations suggest that levels of synaptically released GABA from the terminals of SCN output neurons can influence the relative contribution of pre- versus postsynaptic GABA B receptors in modulating both excitatory and inhibitory SCN innervation to parvocellular PVN neurons.

Optical imaging of long-lasting depolarization on burst stimulation in area CA1 of rat hippocampal slices.

Postsynaptic depolarization of dendrites paired with spike generation at the soma is considered to be a central mechanism of long-term potentiation (LTP) induction and a prime example of a Hebbian synapse. This pairing, however, has never been actually demonstrated on tetanic stimulation. Optical imaging of neural activity with a voltage-sensitive dye (VSD) is one potentially suitable method for examining this pairing. It is possible with optical recording to examine simultaneously the excitation of postsynaptic neurons at multiple sites. Thus the pairing of spike generation at the soma and dendritic depolarization can be examined with population level optical recording in highly laminar structures such as the hippocampal slice preparation. For example, one can correlate the optical signals obtained from cell layers with the activity of the soma, and, similarly, optical signals from stratum radiatum can be correlated with the activity of the apical dendrite, even though one cannot calibrate the optical signals in terms of actual membrane potential. Using the VSD aminonaphthylethenylpyridinium in rat hippocampal slices, we aimed to examine the pairing. Standard tetanic stimulation (100 Hz, 1 s) that elicited LTP in the field excitatory postsynaptic potential (fEPSP) resulted in a long-lasting depolarizing optical signal (about 2 s) that spread progressively along the known input pathway of CA1. The time course of this long-lasting depolarization was similar to that recorded intracellularly and to that reflected in the fEPSP. The long-lasting depolarization was insensitive to D,L-2-amino-5-phosphonovaleric acid (D,L-APV, 50 microM), but D,L-APV inhibited the induction of LTP; this allowed us to increase the signal-to-noise ratio of the optical signal by averaging several trials. Using this improved optical signal, we confirmed that postsynaptic cells practically "missed" spikes during tetanic stimulation in most parts of CA1, which had been suggested in the intracellular recordings. Intracellular recordings revealed a 23% reduction in input resistance, which might explain the failed spike generation at the soma via shunting. A steep spatial convergence of the depolarization along the transverse axis of area CA1 was observed. In contrast to the response resulting from a standard 100-Hz tetanus, broader activation, and paired depolarization with somatic spikes was observed on theta-burst stimulation. Overall we concluded that postsynaptic spike generation, at least in synchronous form, has less effect on LTP induction with standard tetanic stimulation, while theta-burst tetanic stimulation can elicit pairing of dendritic depolarization and somatic discharge.

ATP as a Putative Sensory Mediator: Activation of Intrinsic Sensory Neurons of the Myenteric Plexus via P2X Receptors

The mucosal terminals of sensory neurons intrinsic to the wall of the intestine are sensitive to the chemical environment within the lumen. Lumenal stimuli probably release sensory mediators from the mucosal epithelium, which then activate the nerve terminals indirectly. Here, we tested the idea that ATP activates intrinsic sensory nerve terminals in a way consistent with its being a sensory mediator. We made intracellular recordings from intrinsic sensory neurons located in the myenteric plexus [identified as AH neurons, which are neurons with a long-lasting afterhyperpolarization following the action potential (AP)], located within 1 mm of intact mucosa. Focal electrical stimulation of the mucosa was used to locate and map regions innervated by each neuron. Application of ATP (1-2 mm in the pressure pipette) to these regions elicited trains of APs that originated at the sensory terminals. ATP-gamma-S produced a similar response, but alpha,beta-methylene ATP and 2-methylthio-ATP were only weakly active. The P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',5'-disulphonic acid (PPADS) (60 microm in the bath) abolished the APs evoked by ATP and ATP-gamma-S but spared similar responses evoked by 5-hydroxytryptamine (5-HT). Another P2 receptor antagonist suramin (100 microm in the bath) did not significantly change the number of APs evoked by ATP. Either ATP or alpha,beta-methylene ATP desensitized the ATP-evoked APs; 50% recovery occurred after approximately 5 sec. The number of APs evoked by ATP was reduced, but not abolished, by the selective 5-HT3 receptor antagonist granisetron (1 microm in the bath). ATP was applied to the cell bodies of sensory neurons to investigate whether the cell bodies express the same P2X receptor as the terminals. ATP evoked a fast depolarization associated with a reduction in input resistance and a reversal potential of -11 mV. This depolarization was potentiated by suramin and blocked by PPADS. We conclude that activation of an atypical excitatory P2X receptor by ATP triggers AP generation in the mucosal processes of the sensory neurons; endogenous 5-HT release may also contribute to activation of the nerve terminals. A similar P2X receptor exists on the cell body of the sensory neuron. Together, these data are consistent with a role for ATP as a sensory mediator in gastrointestinal chemosensory transduction.

Biophysical properties and responses to glutamate receptor agonists of identified subpopulations of rat geniculate ganglion neurons

The goal of the current study was to evaluate the electrophysiological properties and responses to glutamate receptor agonists of rat geniculate ganglion (GG) neurons innervating the tongue. Subpopulations of GG neurons were labeled by injecting Fluoro-Gold (FG) or True Blue chloride into the anterior tongue and soft palate (AT and SP neurons) and applying FG crystals to the posterior auricular branch of the facial nerve (PA neurons). Three to 12 days later, the GG neurons were acutely isolated and patch clamped. Although many biophysical properties of the AT, SP and PA neurons were similar, significant differences were found among these groups in properties related to cell excitability. For example, the average amount of current necessary to elicit an action potential was 61 pA in AT neurons ( n=55), 90 pA in SP neurons ( n=41) and 189 pA in PA neurons ( n=35, P<0.001). In addition, AT neurons tended to fire significantly more action potentials during depolarization as well as following hyperpolarizing pulses than SP or PA neuron types. Most GG neurons responded to application of glutamate receptor agonists. The neurons responded with a depolarization accompanied by a reduction in input resistance. These results suggest that subpopulations of neurons in the geniculate ganglion have distinct biophysical properties and express functional glutamate receptors. The differing biophysical properties of GG neurons is possibly related to their functional heterogeneity and glutaminergic neurotransmission may function in the processing of gustatory, and other sensory information, within the geniculate ganglion and its projections.

GABAergic Inhibition in Nucleus Magnocellularis: Implications for Phase Locking in the Avian Auditory Brainstem

In the avian auditory brainstem, nucleus magnocellularis (NM) functions to relay phase-locked signals to nucleus laminaris for binaural coincidence detection. Although many studies have revealed that NM neurons exhibit intrinsic physiological and anatomical specializations for this purpose, the role of inhibition has not been fully explored. The present study characterizes the organization of GABAergic feedback to NM. Anterograde and retrograde labeling methods showed that NM receives a prominent projection from the ipsilateral superior olivary nucleus (SON). The functional features of this projection were explored in a brain slice preparation. Stimulating fibers from the SON evoked long-lasting, depolarizing responses in NM neurons that were blockable by bicuculline, a GABA(A) receptor antagonist. The slow time course of these responses allowed them to undergo temporal summation during repetitive stimulation. The summed GABAergic response was capable of blocking spikes generated in NM neurons by suprathreshold current injection. This inhibitory effect was attributable to a large reduction in input resistance caused by a combination of the opening of a GABAergic Cl(-) conductance and the recruitment of a low-voltage activated K(+) conductance. This large reduction of input resistance increased the amount of current necessary to drive NM neurons to threshold. The results lead us to propose that GABAergic inhibition enhances phase-locking fidelity of NM neurons, which is essential to binaural coincidence detection in nucleus laminaris.

Activation of rat locus coeruleus neuron GABA A receptors by propofol and its potentiation by pentobarbital or alphaxalone

The action of propofol on the rat locus coeruleus was examined using intracellular recording from in vitro brain slice preparations. Concentrations of propofol between 3 and 300 μM were tested. At 100 μM, propofol completely inhibited the firing of all neurons tested ( n=34); this was associated with a 5.7-mV hyperpolarization (range 0–16 mV, n=33) and a 35.6% reduction in input resistance (range 7.3–66.1%, n=33). The propofol-induced responses were not affected by 2-hydroxysaclofen (50 μM) or BaCl 2 (300 μM), but were completely blocked by bicuculline methiodide (100 μM) or picrotoxin (100 μM), indicating that propofol acts on GABA A receptors. As assessed by inhibition of the spontaneous firing rate, propofol was 5.6-fold more potent than GABA (γ-aminobutyric acid). Potentiation of the propofol effect by other general anesthetics or other drugs was also investigated. When pentobarbital (100 μM) was tested alone on locus coeruleus cells, no change in membrane potential or input resistance was seen and there was only a 20.3±7.2% ( n=8) inhibition of firing rate; however, in combination with 30 μM propofol, it caused a 6.1-fold greater increase in membrane hyperpolarization and a 9.7-fold greater reduction in input resistance than 30 μM propofol alone. A relatively low concentration of alphaxalone (10 μM), when tested alone, had little effect on the membrane potential or input resistance and only produced a 46.0±8.9% ( n=8) inhibition of firing rate; however, in combination with 30 μM propofol, it caused a 9.3-fold greater hyperpolarization and an 8.6-fold greater reduction in input resistance compared with 30 μM propofol alone. In contrast, diazepam caused no potentiation of either propofol- or GABA-induced responses. Our data also indicate that locus coeruleus neuron GABA A receptors possess distinctive pharmacologic characteristics, such as blocking of the propofol effects by zinc and insensitivity to diazepam and the direct action of pentobarbital. On the basis of these pharmacologic properties, we suggest that locus coeruleus neuron GABA A receptors do not contain the γ subunit.

Structure–activity relationships of naturally occurring and synthetic opioid tetrapeptides acting on locus coeruleus neurons

Intracellular recording was used to study the effects of eight opioid tetrapeptides with similar amino acid sequences, namely endomorphin-1 ( Tyr-Pro -Trp-Phe-NH 2), endomorphin-2 ( Tyr-Pro -Phe-Phe-NH 2), morphiceptin ( Tyr-Pro -Phe-Pro-NH 2), hemorphin-4 ( Tyr-Pro -Trp-Thr), Tyr-MIF-1 ( Tyr-Pro -Leu-Gly-NH 2), Tyr-W-MIF-1 ( Tyr-Pro -Trp-Gly-NH 2), TAPS ( Tyr- d-Arg -Phe-Sar) and DALDA ( Tyr- d-Arg -Phe-Lys-NH 2), on neurons of the rat locus coeruleus, using a submerged brain slice preparation. All the tetrapeptides inhibited the spontaneous firing of all neurons of the locus coeruleus tested. Higher concentrations also caused hyperpolarization of the neurons and a reduction in input resistance. These inhibitory effects were rapidly and completely reversed by CTAP ( d-Phe-Cys-Tyr- d-Trp-Arg-Thr-Pen-Thr-NH 2, a selective μ-opioid receptor antagonist). The IC 50 of the opioid tetrapeptides, in terms of inhibition of spontaneous firing of locus coeruleus neurons, as compared to the concentrations which produced a 5-mV hyperpolarization (HC 5 mV) were calculated, giving the same rank order of potency: TAPS (IC 50=1.9 nM, HC 5 mV=3.4 nM)>endomorphin-1 (IC 50=8.8 nM, HC 5 mV=22.1 nM) and endomorphin-2 (IC 50=5.3 nM, HC 5 mV=16.1 nM)>DALDA (IC 50=20 nM, HC 5 mV=143 nM)>morphiceptin (IC 50=65 nM, HC 5 mV=335 nM)>Tyr-W-MIF-1 (IC 50=3.8 μM, HC 5 mV=6.7 μM)>hemorphin-4 (IC 50=6.7 μM, HC 5 mV=36.9 μM)>Tyr-MIF-1 (IC 50=37.5 μM, HC 5 mV=76.2 μM). Comparison of the ability of endomorphin-1 and endomorphin-2 to inhibit spontaneous firing based on single-cell recordings ( n=5) showed these two peptides to be equipotent. Based on these results, the structure–activity relationships of these opioid tetrapeptides are discussed herein.

Effects of GABA on acutely isolated neurons from the gustatory zone of the rat nucleus of the solitary tract.

Responses of acutely isolated neurons from the rostral nucleus of the solitary tract (rNST) to GABA receptor agonists and antagonists were investigated using whole-cell recording in current clamp mode. The isolated neurons retain their morphology and can be divided into multipolar, elongate and ovoid cell types. Most rNST neurons (97%), including all three cell types, respond to GABA with membrane hyperpolarization and a reduction in input resistance. The GABA(A) receptor agonist muscimol reduces neuronal input resistance in a concentration-dependent manner, whereas the GABA(B) receptor agonist baclofen had no effect on any of the neurons tested. The GABA and muscimol reversal potentials were both found to be -75 mV Both the GABA competitive antagonist picrotoxin and the GABA(A) receptor antagonist bicuculline block the effect of GABA in a concentration-dependent manner. These results suggest that GABA activates all neurons in the rNST and that inhibitory synaptic activity is important in brainstem processing of gustatory and somatosensory information.

Effects of GABA on neurons of the gustatory and visceral zones of the parabrachial nucleus in rats

Response characteristics of neurons in the gustatory and visceral zone of the parabrachial nucleus (PBN) to γ-aminobutyric acid (GABA) were examined using whole cell recordings in brain slices of the rat. Based on the recording site, neurons were divided into three groups: neurons in the dorsolateral quadrant of the PBN (DL-neurons), neurons in the dorsomedial quadrant of the PBN (DM-neurons) and neurons in the ventromedial quadrant of the PBN (VM-neurons). Recordings were made from 44 DL-, 43 DM-, 39 VM-neurons. Superfusion of GABA resulted in a concentration-dependent reduction in input resistance in 67.5% of the neurons in the PBN (73.1% of the DL-, 62.5% of the DM-, 66.7% of the VM-neurons). No obvious difference of the concentration–response curve was found among three groups. The mean reversal potential of the GABA effect was about −74 mV and no significant differences were observed among three groups of neurons. The GABA response was partly or completely blocked by the GABA A antagonist bicuculline in all neurons tested. Superfusion of the GABA A agonist muscimol resulted in a decrease of the input resistance in all neurons tested. It was concluded that GABA functions as an inhibitory neurotransmitter in both gustatory and visceral part of the PBN, mediated in part, by GABA A receptors.

Serotonin inhibits epileptiform discharge by activation of 5-HT 1A receptors in CA1 pyramidal neurons

The anti-epileptiform effect of serotonin was characterized in cellular models of epilepsy using electrophysiological recording techniques. In the bicuculline model, both serotonin (20 μM) and its 5-HT 1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT, 10 μM) completely blocked the epileptiform discharge and caused membrane hyperpolarization and reduction in input resistance. These effects were completely antagonized by the 5-HT 1A receptor antagonist N-t-butyl-3(4-[2-methoxyphenyl]piperazin-1-yl)-2-phenyl-propanamide (WAY 100135) (10 μM). Epileptiform discharge induced by positive current injection was also blocked by serotonin. The presence of WAY 100135 renders serotonin ineffective in the same model. In the bicuculline model, epileptiform discharge blocked by serotonin reappeared and was also intensified when BaCl 2 was added to the medium. To rule out the possibility of serotonin-induced hyperpolarization strengthening the inhibitory effect of endogenous Mg 2+ on glutamate N-methyl-D-aspartic acid (NMDA) receptor we studied the antiepileptic effect of serotonin in the 0 Mg 2+ model. Spontaneous activity and evoked bursts seen with the OMg 2+ model were completely blocked by serotonin. WAY 100135 completely antagonized serotonin effects in this model as well. This study provides evidence suggesting that in rat CA1 pyramidal neurons, serotonin can inhibit epileptiform activity in a variety of accepted epilepsy cellular models and that inhibition of epileptiform bursts by serotonin may be mediated by activation of the 5-HT 1A receptor subtype.

Regulation of muscarinic acetylcholine receptor-mediated synaptic responses by adenosine receptors in the rat hippocampus.

1. Intracellular current clamp recordings were made from CA1 pyramidal neurones in rat hippocampal slices. Experiments were performed in the presence of ionotropic glutamate receptor antagonists and gamma-aminobutyric acid (GABA) receptor antagonists to block all fast excitatory and inhibitory synaptic transmission. A single stimulus, delivered extracellularly in the stratum oriens, caused a reduction in spike frequency adaptation in response to a depolarizing current step delivered 2 s after the stimulus. A 2- to 10-fold increase in stimulus intensity evoked a slow excitatory postsynaptic potential (EPSP) which was associated with a small increase in input resistance. The peak amplitude of the EPSP occurred approximately 2.5 s after the stimulus and its magnitude (up to 30 mV) and duration (10-50 s) increased with increasing stimulus intensity. 2. The slow EPSP was unaffected by the metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG; 1000 microM) but was greatly enhanced by the acetylcholinesterase inhibitor physostigmine (1-5 microM). Both the slow EPSP and the stimulus-evoked reduction in spike frequency adaptation were inhibited by the muscarinic acetylcholine receptor (mAChR) antagonist atropine (1-5 microM). These results are consistent with these effects being mediated by mAChRs. 3. Both the mAChR-mediated EPSP (EPSPm) and the associated reduction in spike frequency adaptation were reversibly depressed (up to 97%) by either adenosine (100 microM) or its non-hydrolysable analogue 2-chloroadenosine (CADO; 0.1-5.0 microM). These effects were often accompanied by postsynaptic hyperpolarization (up to 8 mV) and a reduction in input resistance (up to 11%). The selective adenosine A1 receptor agonists 2-chloro-N6-cyclopentyladenosine (CCPA; 0.1-0.4 microM) and R(-)N6-(2-phenylisopropyl)-adenosine (R-PIA; 1 microM) both depressed the EPSPm. In contrast, the adenosine A2A receptor agonist 2-p-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS 21680; 0.5-1.0 microM) did not significantly affect the EPSPm. 4. The selective adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 0.2 microM) fully reversed the depressant effects of both adenosine (100 microM) and CADO (1 microM) on the EPSPm and the stimulus-evoked reductions in spike frequency adaptation. 5. DPCPX (0.2 microM) alone caused a small but variable mean increase in the EPSPm of 22 +/- 19% and enabled activation of an EPSPm by a previously subthreshold stimulus. In contrast, the selective adenosine kinase inhibitor 5-iodotubercidin (5-IT; 10 microM) inhibited the EPSPm by 74 +/- 10%, an effect that was reversed by DPCPX. 6. The concentration-response relationship for the depressant action of CADO on the EPSPm more closely paralleled that for its presynaptic depressant action on glutamate-mediated EPSPs than that for postsynaptic hyperpolarization. The respective mean IC50 and EC50 concentrations for these effects were 0.3, 0.8 and 3.0 microM. 7. CADO (1-5 microM) did not have a significant effect on the postsynaptic depolarization, increase in input resistance and reduction in spike frequency adaptation evoked by carbachol (0.5-3.0 microM). All these effects were abolished by atropine (1 microM). 8. These data provide good evidence for an adenosine A1 receptor-mediated inhibition of mAChR-mediated synaptic responses in hippocampal CA1 pyramidal neurones. This inhibition is mediated predominantly presynaptically, is active tonically and can be enhanced when extracellular levels of endogenous adenosine are raised.

Serotonin-induced modulation of excitability in an identified Helisoma trivolvis neuron

Exogenous serotonin (5-hydroxytryptamine; 5-HT) induced inhibitory effects in Helisoma trivolvis buccal neuron 5 (B5), which included a hyperpolarization of the membrane potential, a reduction in input resistance and a decrease in the stimulus-evoked spiking rate. The reversal potential for the 5-HT-induced hyperpolarization was -88.7 mV, suggesting the involvement of an enhanced K+ conductance. Input/output curves measuring the number of action potentials evoked by current injection demonstrated a 5-HT-induced reduction in B5 neuronal excitability that was dose-dependent with a 50 % inhibitory concentration (IC50) of 1.1 &micro;mol l-1. These inhibitory effects on neuron B5 were in striking contrast to the excitatory effects of this neurotransmitter on the buccal radular tensor motoneuron B19. In this neuron, 5-HT elicited a sustained depolarization and enhanced spiking activity. Previous cell culture studies have demonstrated that 5-HT also inhibits B19 growth cone motility and neurite outgrowth, but has no effect on the extension of neuritic processes from neuron B5. These effects, and the divergent effects of 5-HT on the outgrowth of neurons B5 and B19, probably result from differential effects of 5-HT on neuronal membrane potential and excitability. Modulation of the levels of spiking activity, translated into changes in neuronal Ca2+ levels, would cause alterations in growth cone motility. Previous studies have shown that the effects of 5-HT on B19 membrane potential and neurite extension are cyclic-AMP-dependent. We have used specific activators and inhibitors of signal transduction mechanisms to demonstrate that the 5-HT-induced reductions in B5 neuronal excitability are cyclic-AMP-independent and may involve the lipoxygenase pathway of arachidonic acid metabolism. Our results support the hypothesis that modulatory signals and signal transduction pathways regulating the plasticity of mature neural circuits may also modulate the differentiation of the neurons comprising those networks during development.

Effects of neurochemically excited group III–IV muscle afferents on motoneuron afterhyperpolarization

When humans voluntarily and maximally contract a muscle under isometric conditions, the average firing rate of motor units decreases from an initially high value over several tens of seconds. The mechanisms underlying the rate reduction are probably manifold. One mechanism could involve changes in the motoneuron afterhyperpolarization, and another reflex effects of group III–IV muscle afferents that are excited during developing muscle fatigue. It appears possible that changes in motoneuron afterhyperpolarization are mediated by these afferent inputs. We therefore studied effects on motoneuron afterhyperpolarization of small-diameter muscle afferents excited by intra-arterially injected metabolites such as bradykinin and serotonin. In decerebrate and mostly spinalized cats, lumbosacral α-motoneurons were recorded intracellularly. Current pulses were injected to test for input resistance and elicit action potentials and afterhyperpolarizations. Afterhyperpolarizations were averaged from 10 successive stimulus repetitions. Measurements were taken of afterhyperpolarization amplitude, half-width and area; and exponential functions were fitted to the afterhyperpolarization decay phase to determine afterhyperpolarization decay time-constants. In selected cases, the entire afterhyperpolarization trajectory was fitted with a sum of two exponentials to assess more precisely changes in afterhyperpolarization trajectory. Small catheters were inserted into side-branches of the sural artery and the accompanying vein to apply substances like bradykinin, serotonin and KCl to the calf muscles. Concentrations were in the range of those used by other workers. Intra-arterial injection of bradykinin and serotonin usually decreased blood pressure, which may at times have affected mean motoneuron membrane potentials. Afterhyperpolarization amplitude usually changed with membrane potential in a way expected from ensuing changes in driving potential. Whenever excitation of group III–IV muscle afferents caused moderate to strong increases in motoneuron synaptic noise, afterhyperpolarization amplitudes were reduced, usually in parallel to decreases in input resistance. Afterhyperpolarization half-widths were mostly unaffected, but occasionally decreased. There was a significant trend for afterhyperpolarization decay time-constants to increase during increased synaptic noise, this increase being inversely correlated with the reduction in afterhyperpolarization amplitude. The reduction in input resistance was associated with a decrease in the membrane time-constant, which could therefore not account for the prolongation of the afterhyperpolarization decay time-constant. The afterhyperpolarization area decreased, indicating that the reduction of afterhyperpolarization amplitude outweighed the prolongation of afterhyperpolarization decay time-constant.During a prolonged fatiguing muscle contraction group III–IV afferents become increasingly excited, produce augmenting synaptic inputs in motoneurons, and will change afterhyperpolarization properties. On average, these changes per se tend to diminish the effect of afterhyperpolarization on motoneuron discharge.

Neuronal diversity in the subiculum: correlations with the effects of somatostatin on intrinsic properties and on GABA-mediated IPSPs in vitro.

1. We used intracellular current-clamp techniques to record from 33 ventral subicular neurons in slices or rat hippocampal formation. Presumed pyramidal neurons were characterized by their responses to depolarizing current pulses as either intrinsically burst firing (IB) or regular spiking (RS). Within the subiculum, IB cells were encountered most frequently in the deep cell layer, whereas RS cells were encountered most frequently in the superficial cell layer. IB cells had more depolarized resting potentials, lower input resistances, and more sag in their voltage responses to hyperpolarizing current pulses. 2. Somatostatin (5 microM) applied in the bathing medium caused a hyperpolarization and reduction in input resistance. These effects were of greater magnitude in IB cells. Somatostatin had no effect on sag in either cell type. These effects of somatostatin were unchanged in the presence of gamma-aminobutyric acid (GABA) receptor antagonists. 3. In a series of experiments conducted in RS cells only, somatostatin reduced the amplitude of the late but not the early component of evoked biphasic inhibitory postsynaptic potentials (IPSPs). 4. A second series of experiments was conducted in RS and IB cells. Somatostatin reduced the amplitude of pharmacologically isolated GABAA IPSPS in both cell types. In IB cells but not RS cells there was a correlation between this effect and the somatostatin-induced hyperpolarization. Somatostatin also reduced the amplitude of isolated GABAB IPSPS in both cell types, but more so in IB cells. 5. Somatostatin had no effect on the reversal potential of either IPSP in either cell type and no effect on the GABAA-mediated conductance in either cell type. In contrast, the GABAB-mediated conductance was reduced, especially in IB cells. 6. The effects of somatostatin on GABAA IPSPS are principally a result of membrane shunting and reductions in ionic driving force, but these mechanisms do not account for the reduction in GABAB IPSPS. 7. We suggest that the combined effects of somatostatin are likely to alter the balance between fast and slow inhibition and to do so more in IB cells than in RS cells.

2324 A method to estimate the real [Ca 2+] i from signals of fluorescentcalcium dyes

1.1. Neuron LP1 of Hermissenda crassicornis shows pronounced anomalous rectification at a membrane potential of −65 mV, or more negative, indicated by a 3- to 4-fold reduction in input resistance, producing an S-shaped current-voltage relationship.2.2. Axotomized LPls show membrane potential bistability since shifts in membrane potential between stable levels at ca −45 and −65 mV accompanied by a change in input resistance occur spontaneously or are induced by current pulses.3.3. Stability at −65 mV appears to be due to activation of the anomalous rectifier.

Action of dexmedetomidine on rat locus coeruleus neurones: intracellular recording in vitro

The action of dexmedetomidine on rat locus coeruleus neurones was examined using intracellular recordings from the in vitro brain slice preparation. Concentrations of dexmedetomidine from 1 to 1000 nM were tested. At 30 nM, dexmedetomidine produced complete inhibition of firing of all neurones tested ( n = 21); this was associated with a 13 mV hyperpolarization (range 2.2–29.7 mV, n = 21) and a 27% reduction in input resistance (range 11.1–46.2%, n = 17). The dexmedetomidine responses reached a plateau phase between 100 and 1000 nM. Based on single-cell recordings, the hyperpolarizing potency of dexmedetomidine was found to be 6 times greater than that of clonidine ( n = 10). The reversal potential for the dexmedetomidine-induced hyperpolarization was − 106.9 ± 1.7 mV ( n = 9), a value similar to the K + equilibrium potential; hyperpolarization was blocked by both CsCl and BaCl 2. The effect of dexmedetomidine was antagonized by yohimbine, with a dissociation equilibrium constant of 30 nM. In contrast, prazosin, the α 1-, α 2B- and α 2C-adrenoceptor subtype-preferring ligand, did not inhibit the dexmedetomidine effect. Our results also show that low concentrations of oxymetazoline (10–300 nM), an α 2A-adrenoceptor subtype-selective drug, cause profound inhibition of neuronal activity in the locus coeruleus. These data therefore suggest that dexmedetomidine binds to α 2A-adrenoceptors on the cell membrane of neurones of the locus coeruleus and that this leads to opening of the inwardly rectifying K + channels, resulting in the observed hyperpolarization of the membrane.

Internal Ca2+ stores involved in anoxic responses of rat hippocampal neurons.

1. During whole-cell recordings from CA1 neurons of rat brain slices with electrodes containing only KMeSO4 and Hepes, brief anoxia (2-3 min) consistently evoked a hyperpolarization (delta V approximately 14 mV) and reduction in input resistance (delta R approximately -20%). 2. As in previous intracellular recordings, Dantrolene sodium (10 microM) suppressed the anoxic delta V and delta R, confirming the release of internal Ca2+ is a major component of the anoxic response. 3. To identify the relevant intracellular Ca2+ store, other blockers of Ca2+ release were applied either externally (in the bath) or internally, by addition to the contents of the recording electrode. 4. The anoxic hyperpolarization was abolished or much reduced by heparin (10-20 micrograms ml-1, internal), thapsigargin (10 microM, external), Ruthenium Red (50 microM, internal) and external procaine (0.5-2 mM), but not by internal procaine (0.5-1 mM) or ryanodine (10 microM, external). 5. The anoxic fall in resistance was also abolished or reduced by heparin, thapsigargin and external procaine, but not by ryanodine, internal procaine or Ruthenium Red. 6. In addition, external procaine (0.5-2 mM) eliminated the early (transient) depolarization and reduced the post-anoxic hyperpolarization by 60 +/- 22%. 7. None of these agents consistently changed the resting potential, but the input resistance was significantly increased by Dantrolene and external procaine. 8. In view of the marked effects of heparin and thapsigargin, but not ryanodine and internal procaine, we conclude that the anoxic response seen in such whole-cell recordings is initiated predominantly by Ca2+ release from an internal store that is InsP3 sensitive rather than Ca2+ sensitive. 9. Comparable but less pronounced effects of external procaine were seen during intracellular recordings with 3 M KCl-containing electrodes. The dose-dependent suppression of various features of the anoxic response by external procaine (EC50 approximately 0.2 mM) is presumed to be mediated by a superficial membrane trigger or modulating site.

Effects of muscarinic receptor stimulation of sympathetic preganglionic neurones of neonatal rat spinal cord in vitro

Whole cell current-clamp recordings were made from 85 sympathetic preganglionic neurones (SPN) of the neonatal spinal cord in vitro. Superfusion of up to 500 μM acetylcholine (2–30 sec) gave weak responses. Carbachol (CChol; 5–50 μM) superfused for 1–20 sec, hyperpolarized SPN. This response was associated with a mean reduction in input resistance of 22%. Ion substitution studies suggested that potassium is the likely carrier of at least part of the current underlying the CChol-induced hyperpolarization. Some SPN display spontaneous rhythmic oscillations in their membrane potentials which may be due to action potential discharge in electrically-coupled neurones. CChol also acts to inhibit these oscillations concomitant with the hyperpolarization. Responses to CChol were unaffected by addition of 500 nM TTX to the bathing medium suggesting that CChol acts directly upon SPN. Carbachol-induced hyperpolarizing responses were totally abolished by the non-specific muscarinic receptor antagonist atropine (20–50 μM). Pirenzepine, at concentrations over 5 μM reversibly reduced the responses to CChol. Gallamine, an M2 receptor antagonist applied at 25 μM also reversibly abolished the CChol responses. These results suggest that CChol-mediated hyperpolarizations may be due to M2 receptor activation.

Cholinergic synaptic transmission between proprioceptive afferents and a hind leg motor neuron in the locust.

1. The physiology and pharmacology of the connections made by stress-sensitive campaniform sensilla on the proximal dorsal tibia of a locust hind leg with a fast extensor tibiae motor neuron (FETi) have been examined. 2. Mechanical stimulation of a campaniform sensillum on the anterior surface of the tibia elicited bursts of spikes in its afferent and a depolarization in FETi. Each afferent spike was followed at short and constant latency by an excitatory postsynaptic potential (EPSP) in FETi, even at high-frequency stimulation. Electrical stimulation of the extensor tibiae muscle with the tibia fixed elicited an antidromic spike in FETi followed by a compound potential which resulted from the activation of the campaniform sensillum afferent. The connections between the campaniform sensillum and FETi was monosynaptic and chemically mediated. 3. Ionophoresis of ACh into the neuropil depolarized FETi, as did the application of the cholinesterase inhibitor physostigmine. Bath application of the cholinergic agonists, nicotine and muscarine, also depolarized FETi, with nicotine causing a reduction in input resistance, while muscarine caused no detectable change in input resistance. Muscarine and the muscarinic agonist oxotremorine also caused rhythmic depolarizations and bursts of spikes in FETi. These effects were seen in low calcium/high magnesium saline to block synaptic transmission and are therefore due to direct effects on FETi. FETi therefore appears to have both nicotinic and muscarinic receptors. 4. The compound potential in FETi, caused by activation of the campaniform sensillum, was reduced by bath application of nicotinic cholinergic antagonists. However, the muscarinic antagonist scopolamine increased the amplitude of the compound potential.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of temperature on properties of flight neurons in the locust

High ambient temperatures increase the wing-beat frequency in flying locusts, Locusta migratoria. We investigated parameters of circuit and cellular properties of flight motoneurons at temperatures permissive for flight (20–40 °C). As the thoracic temperature increased motoneuronal conduction velocity increased from an average of 4.40 m/s at 25 °C to 6.73 m/s at 35 °C, and the membrane time constant decreased from 11.45 ms to 7.52 ms. These property changes may increase locust wing-beat frequency by affecting the temporal summation of inputs to flight neurons in the central circuitry. Increases in thoracic temperature from 25–35 °C also resulted in a hyperpolarization of the resting membrane potentials of flight motoneurons from an average of-41.1 mV to -47.5 mV, and a decrease of input resistances from an average of 3.45 MΩ to 2.00 MΩ. Temperature affected the measured input resistance both by affecting membrane properties, and by altering synaptic input. We suggest that the increase in conduction velocity Q10=1.53) and the decrease of membrane time constant (Q10=0.62) would more than account for the wing-beat frequency increase (Q10=1.15). Hyperpolarization of the resting membrane potential (Q10=1.18) and reduction in input resistance (Q10=0.54) may be involved in automatic compensation of temperature effects.