Post-stimulus Hyperpolarization Research Articles

In vivo 4-aminopyridine treatment alters the neurotoxin 3-acetylpyridine-induced plastic changes in intrinsic electrophysiological properties of rat cerebellar Purkinje neurones

Electrophysiological dysfunction of Purkinje cells causes cerebellar ataxia. Recent studies indicated that 4-aminopyridine (4-AP) can prevent the attacks in patients with episodic ataxia type 2. However, the cellular mechanism(s) by which 4-AP might be beneficial for the improvement of motor function remain unclear. Here, electrophysiological and behavioural consequences of in vivo co-treatment with 4-AP against 3-acetylpyridine (3-AP)-induced ataxia in rats were assessed. Combined treatment with 4-AP partially improved motor behaviour compared to the ataxic rats. Treatment with 3-AP alone induced plastic alterations in the cells' intrinsic properties, so that the latency of the initial neural spike was significantly increased ( P < 0.001); however, both instantaneous firing frequency and amplitude of calcium spikes were significantly ( P < 0.001) suppressed. 3-AP treatment also resulted in significant decrease in the duration of action potential ( P < 0.05) and the amplitude of afterhyperpolarization (( P < 0.05) as well as post-stimulus hyperpolarization potentials ( P < 0.001). Purkinje cells in rats co-treated with 4-AP, however, fired predominantly in rhythmic bursts. The mean amplitude of Ca 2+ spikes was significantly ( P < 0.001) greater compared to ataxic rats, but similar to control value. As evidenced by a significant decrease ( P < 0.001) in the first spike latency, the cells' intrinsic excitability was also increased. In 4-AP co-treated group, the duration of action potential was also significantly lengthened ( P < 0.001) compared to control and 3-AP group. These results suggest that modulation of intrinsic electrical properties and potentiation of Ca 2+ channels function caused by in vivo 4-AP treatment is likely to be partly responsible for its neuroprotective action.

Phenotypes of trpl Mutants and Interactions between the Transient Receptor Potential (TRP) and TRP-Like Channels inDrosophila

The trp and trpl genes are thought to encode two classes of light-activated ion channels in Drosophila. A previous report indicated that a null trpl mutant does not display any mutant phenotype. This lack of detectable mutant phenotypes made it difficult to suggest functions for the transient receptor potential-like (TRPL) channel in photoreceptor responses. Here, the properties of trpl photoreceptor responses were studied by using electroretinogram (ERG) and intracellular recording techniques in combination with light stimuli of relatively long durations. Distinct mutant phenotypes were detectable under these conditions. These consisted of a reduced sustained component, oscillations superimposed on the response, a poststimulus hyperpolarization, and altered adaptation properties to dim background light. Comparison of photoreceptor responses obtained from wild type, trp, and trpl showed that the responses obtained from the trp and trpl null mutants did not sum up to that of the wild-type response. To explain the nonlinear summation at the peak of the response, Reuss et al. (1997) proposed that Ca(2+) ions entering through the TRP channel modulate TRP and TRPL channel activities differentially. However, nonlinear summation was present not only at the peak but throughout the duration of response. Two lines of evidence are presented to suggest that, in addition to the interaction proposed by Reuss et al. (1997), there are other forms of interactions between TRP and TRPL channels, probably involving the channel proteins themselves.

In Vivo Ca 2+ Dynamics in a Cricket Auditory Neuron: An Example of Chemical Computation

Fura-2 calcium imaging in the cricket omega neuron revealed increased intracellular free calcium ion concentration in response to simulated cricket calling songs and other sound stimuli. The time course of the increase and decrease in intracellular calcium coincided with the time course of forward masking, a time-dependent modulation of auditory sensitivity. The buffering of calcium transients with high concentrations of a kinetically fast calcium buffer eliminated the post-stimulus hyperpolarization associated with forward masking, whereas the uncaging of calcium inside the neuron produced a hyperpolarization. The results suggest that sound-stimulated intracellular calcium accumulation acts by means of a calcium-activated hyperpolarizing current to produce forward masking. These findings underscore the importance of chemical dynamics in neural computation by demonstrating a behaviorally relevant role of calcium dynamics in vivo.

Calcium‐activated hyperpolarizations in rat locus coeruleus neurons in vitro.

1. Intracellular recordings were made from rat locus coeruleus (LC) neurons in completely submerged brain slices. Trains of action potentials in LC neurons were followed by a prolonged post-stimulus hyperpolarization (PSH). If trains were elicited with depolarizing current pulses of sufficient intensity, PSH was composed of a fast, early component (PSHE) and a slow, late component (PSHL). PSH which followed trains elicited with lower intensity depolarizing current pulses consisted only of PSHL. 2. Both PSHE and PSHL were augmented by increasing the number of action potentials in the train and both were associated with an increase in membrane conductance. The reversal potential for PSHE was -108 mV and for PSHL it was -114 mV. 3. When a hybrid voltage clamp protocol was used, the current underlying PSH (IPSH) was observed to consist of an early, rapidly decaying component, IE, followed by a late, slower decaying component, IL. The time course of decay of IPSH was biexponential with the time constant of decay of IL more than one order of magnitude larger than the time constant of decay of IE. An increase in the concentration of external K+ shifted the reversal potentials for IE and IL in the depolarizing direction; the mean value of shift per tenfold increase in external K+ concentration was 57.1 mV for IE and 57.6 mV for IL. 4. Both PSHE and PSHL were inhibited by lowering the external Ca2+ concentration or by application of the Ca2+ channel blockers Cd2+ (200-500 microM) or nifedipine (100 microM). Intracellular injection of EGTA abolished both components of PSH. Increasing the external Ca2+ concentration augmented both PSH components. 5. Superfusion of dantrolene (25 microM) or ryanodine (20 microM) decreased the amplitude and duration of PSHL with much less effect on PSHE. 6. d-Tubocurarine (20-200 microM) selectively blocked PSHE with no effect on PSHL; this effect is the same as that of apamin which we have previously described. Superfusion with charybdotoxin (40 nM) or TEA (400 microM-1 mM) did not reduce PSHE or PSHL. 7. Inhibition of IA by 4-aminopyridine or 2,4-diaminopyridine also did not reduce either component of PSH. In fact, these agents slightly augmented both components of PSH; this effect was probably secondary to the prolongation of action potential duration. Superfusion of TEA in concentrations of 2-10 mM increased the size and duration of PSHL and increased the duration but decreased the size of PSHE.(ABSTRACT TRUNCATED AT 400 WORDS)

In vitro responses of caudal hypothalamic neurons to hypoxia and hypercapnia

Results from previous studies have suggested that the hypothalamus modulates cardiorespiratory responses to hypoxia and/or hypercapnia. Many neurons in the caudal hypothalamus are stimulated by hypercapnia and hypoxia in vivo; however, it is not known if these responses are dependent upon input from other areas. Whole-cell patch and extracellular recordings from a brain slice preparation were used in the present study to determine the direct effects of hypoxia (5% CO2/95% N2 or 10% O2/5% CO2/85% N2) and hypercapnia (7% CO2/93% O2) on caudal hypothalamic neurons in vitro. Coronal sections (400–500 μm) were obtained from young Sprague-Dawley rats and placed in a recording chamber that was perfused with nutrient media equilibrated with 95% O2/5% CO2. Extracellular recordings demonstrated that hypoxia stimulated over 80% of the neurons tested; the magnitude of the response was dependent upon the degree of hypoxia. In addition, over 80% of cells that were excited by hypoxia retained this response during synaptic blockade. Hypercapnia increased the discharge frequency of 22% of the caudal hypothalamic neurons that were studied. A second set of caudal hypothalamic neurons were studied with whole-cell patch recordings; the mean resting membrane potential of these neurons was −51.8 ± 1.0 mV with an average input resistance of 399 ± 49 MΩ. Hypoxia produced a depolarization in 76% of these neurons; a poststimulus hyperpolarization often occurred. A depolarization and/or increase in discharge rate during hypercapnia was observed in 35% of the neurons tested. Only 10% of all neurons studied were excited by both hypoxia and hypercapnia.These findings suggest that separate subpopulations of caudal hypothalamic neurons are sensitive to hypoxia and hypercapnia. Thus, this hypothalamic area may be a site of central hypoxic and hypercapnic chemoreception.

Slowly activating K + channels in rat olfactory receptor neurons

Patch-clamp techniques were used to investigate slowly activating, Ca(2+)-insensitive K+ channels of isolated rat olfactory receptor neurons. These channels had a unitary conductance of 135 pS and were only found in a small proportion (less than 5%) of membrane patches. Upon depolarization to voltages more positive than -50 mV, the channels activated gradually over a period of at least 10 s. When hyperpolarized to negative voltages, channel activity deactivated in a slow but voltage-dependent manner. These channels may underlie a slowly activating K+ current that is observed in approximately 30% of whole-cell recordings. Similar single channels have been reported in smooth muscle cells, but this is the first demonstration of these channels in any type of neuron. The channels may contribute to the spike frequency adaptation and post-stimulus hyperpolarization that are observed during the excitatory response to odorants. They may also contribute to cell repolarization following large odorant-stimulated receptor currents.

Functional significance of the apamin-sensitive conductance in rat locus coeruleus neurons

The effects of apamin on rat locus coeruleus (LC) neurons were studied in a brain slice preparation with intracellular recording. Bath application of apamin (2–500 nM) reduced the amplitude of an intermediate component of the afterhyperpolarization (AHP) following single spontaneous action potentials, but did not change the size or time-course of fast and slow components of the AHP, spike amplitude or duration. Apamin blocked the early component of the post-stimulus hyperpolarization (PSH) which follows a train of action potentials. The size of the late component of PSH was sometimes augmented by apamin. Apamin increased the number of spikes evoked by a depolarizing current pulse and increased the slope of the spike frequency-current intensity relation. Accommodation of firing during long depolarizing pulses showed a biexponential time-course indicating 2 distinct components. Apamin specifically reduced the contribution of the fast component of accommodation and increased its time constant. These data indicate that the apamin-sensitive conductance is functionally important in accommodation at faster firing rates such as those seen during evoked spike trains in the present experiments, and which may occur in vivo during behavioral arousal and in anxiety or drug withdrawal syndromes.

Baclofen increases the potassium conductance of rat locus coeruleus neurons recorded in brain slices

Baclofen causes a concentration-dependent inhibition of spontaneous firing, hyperpolarization and resistance decrease in locus coeruleus (LC) neurons recorded intracellularly in a brain slice preparation. The (−) isomer is active while the (+) isomer has little or no activity which indicates that the baclofen effect is stereoselective. Baclofen action on LC neurons is a direct postsynaptic effect since it remains in low Ca 2+, high Mg 2+ media. Baclofen actions on LC neurons are resistant to the GABA A antagonist bicuculline. The baclofen-induced hyperpolarization reverses at the K + equilibrium potential, as estimated by the reversal potential of the post-stimulus hyperpolarization which follows an evoked train of action potentials. When the K + concentration in the superfusion media is increased, the reversal potential for the baclofen-induced hyperpolarization shifts linearly with a slope of 61 mV per 10-fold change as predicted by the Nernst equation for a pure K + conductance. The baclofen-induced K + conductance increase is prevented by addition of the K +-channel blocker Ba 2+ to the external media. Taken together, these data suggest that baclofen directly hyperpolarizes LC neurons by activation of GABA B receptors which leads to an increase in K + conductance.

Relative contribution of two mechanisms to post-stimulus hyperpolarization in leech retzius cells

1. 1. In Retzius cells of the horse leech, Haemopis sanguisuga, intensive firing is followed by prolonged post-stimulus hyperpolarization (PSH) mediated by an increase in K-conductance and activation of the electrogenic sodium pump. 2. 2. The recovery of membrane potential during PSH can be described by a biexponential model from which the relative contribution of the early and the late component to the peak amplitude of PSH may be calculated. 3. 3. The time course of the calculated early component coincides with the time course of changes that occur in input membrane resistance during PSH. 4. 4. The relative contribution of the late component increases linearly with an increase in stimulus duration. The rate constants of the two phases did not change with changes in stimulus duration. 5. 5. It was concluded that the proposed model is useful for the estimation of the relative contribution and kinetics of two PSH mediating mechanisms (K-conductance increase and activation of an electrogenic sodium pump) in different experimental conditions.

Long-lasting excitability changes of soleus alpha-motoneuron induced by midpontine stimulation in decerebrate, standing cat.

Stimulation of the dorsal portion of the caudal tegmental field (DTF) in the pons resulted in hyperpolarization of extensor alpha-motoneurons (alpha-MNs) that persisted for several minutes after cessation of the stimulation. The resulting inhibition of alpha-MN discharge led to a progressive reduction in the number of active motor units. Renshaw cells, persistently active at high levels of extensor muscle tone, were abruptly silenced by DTF stimulation. Active discharge was renewed at the time of cessation of the stimulation but at a frequency reduced in proportion to the persistently lowered level of extensor muscle tone. Ia primary afferents were tonically active during the high extensor tonus of reflex standing. DTF stimulation was accompanied by a brief, slight increase in Ia discharge frequency followed by a reduction in frequency variably correlated to the magnitude of extensor force reduction. Orthodromically elicited Ia EPSPs in the soleus alpha-MNs were reduced in peak voltage, time to peak, and half width during the hyperpolarization accompanying DTF stimulation. All of these parameters recovered beyond their prestimulus values with the cessation of DTF stimulation in spite of the persisting hyperpolarization. Antidromically initiated invasion of the somatodendritic (SD) segment of the motoneuron membrane was delayed and sometimes blocked during DTF stimulation. At the same time, the peak voltage of the SD action potential was reduced. There was an immediate recovery of these changes on termination of DTF stimulation, although the poststimulus hyperpolarization persisted. Intracellular injection of depolarizing current steps during DTF stimulation revealed a depression of membrane excitability that persisted during the hyperpolarization that followed the termination of the DTF stimulation. Depolarizing and hyperpolarizing steps of intracellular current were used to demonstrate a reduction of cellular input resistance during DTF stimulation. The resistance values rapidly returned to prestimulus levels following the cessation of DTF stimulation. It was demonstrated that the degree of resistance change is greater and that the magnitude of DTF-induced hyperpolarization is smaller for low-resistance cells than for high-resistance cells. Iontophoretically induced increase in intracellular Cl- resulted in a reversal of both Ia IPSPs and the hyperpolarization induced by DTF stimulation. The hyperpolarization enduring after DTF stimulation was not affected by the Cl- injection.

Post-stimulus hyperpolarization in retzius nerve cells of leech, Haemopis sanguisuga

Abstract 1. 1. Post-stimulus hyperpolarization (PSH) that occurred in response to direct or indirect (across the electrotonic junction) depolarizing stimulation has been studied in Retzius neurons of horse leech, Haemopis sanguisuga. 2. 2. The amplitude and time course of PSH were dependent on the duration of depolarizing stimulation and passive membrane properties of Retzius cells. The recovery of membrane potential during PSH has an approximately exponential time course and consisted of two phases: the early (fast) one, which has been associated with a decrease in membrane input and transfer resistance and the late (slow) one. 3. 3. Na-free saline (with TRIS as substituing cation) and K-free saline augmented the amplitude of PSH and abolished its late phase. 4. 4. Ouabain completely blocked the late phase and greatly diminished the early one. Verapamil almost completely abolished PSH. 5. 5. Increased external Ca concentrations slightly augmented the amplitude and duration of PSH. 6. 6. Application of strong inward current reversed the PSH. The PSH reversal potential varied with extracellular K concentrations, approx. 40 mV per ten-fold change in external K+. 7. 7. It was concluded that PSH in Retzius nerve cells is mediated by two mechanisms: the prolonged increase in membrane K conductance and increased activity of the electrogenic sodium pump.

Evidence for A Synaptically Mediated Decrease in Conductance in A Crustacean Myocardium

1. In the neurogenic heart of the isopod Porcellio dilatatus, electrical stimulation of the cardio-regulatory nerves at rates greater than 20-25 pulses/s elicited inhibitory junctional potentials (IJPs) in the myocardium. Its cessation was followed by a long lasting hyperpolarization of myocardial membrane (post-stimulus hyperpolarization = PSH). 2. During the PSH the membrane resistance of the heart muscle increased. The PSH was enhanced by myocardium hyperpolarization, decreased by depolarization and reversed around -50 mV. 3. Picrotoxin inhibited the summated IJPs elicited by the stimulation and thus caused the membrane to maximally hyperpolarize during inhibitory train, thus suggesting a composite nature of the inhibitory processes. 4. The PSH was reversibly reduced in K+-free saline or in ouabain containing saline but partial restoration was obtained by injection of inward current to the myocardium. 5. The PSH was abolished in lithium saline and reduced in Na+-deficient (choline) solution. Cl-deficient solution that markedly affected the summated IJPs shortly after its introduction did not affect the PSH. 6. It is proposed that the PSH results from a decrease in conductance, presumably to both Na+ and K+. The implication of such a mechanism as a component of the inhibitory regulation of this crustacean heart is discussed.

Diphasic synaptic potentials and prolonged poststimulus hyperpolarization in Helix pomatia neurons

Synaptic activity of neurons giving diphasic excitatory-inhibitory potentials in response to orthodromic stimulation was recorded intracellularly. In response to stimulation of nerves by a single short pulse these neurons developed only the excitatory component of the diphasic potential, but with a longer stimulus a prolonged inhibitory phase, partly suppressing the initial excitatory component, was added. The excitatory phase appeared only when the resting potential reached a certain level. In their response to repetitive stimulation, neurons with a diphasic potential are divided into habituating and nonhabituating. The diphasic potential can also arise in response to application of acetylcholine to the soma of these neurons. It is postulated that this potential reflects the response of different receptors of the postsynaptic membrane to the same mediator. Prolonged poststimulus hyperpolarization can be obtained after repetitive orthodromic or direct stimulation of some neurons. However, as analysis of the results showed, poststimulus hyperpolarization is endogenous in origin and differs in its mechanisms from the diphasic potential.

Increased K-conductance as proximate cause of post-stimulus hyperpolarization in Tritonia neurones

1. 1. Post-stimulus hyperpolarization (PSH) up to 10 mV, lasting from 0·5 to 2 min, was observed in neurones of Tritonia following outward-current stimuli. An increase of membrane conductance occurred simultaneously with the PSH. 2. 2. Neither ouabain nor cooling reduced the size of the PSH. 3. 3. By application of inward currents immediately after the outward-current stimulus, a reversal potential was measured for the PSH. The reversal potential varied about 42 mV/tenfold change in external potassium concentration. 4. 4. In the absence of inward current, the PSH increased in size with decreasing external potassium concentration. 5. 5. The results indicate that PSH is not due to increased activity of an electrogenic pump, but results from a long-lasting increase in membrane conductance, primarily to potassium ions.

Post-stimulus hyperpolarization and slow potassium conductance increase in Aplysia giant neurone.

1. Intracellular records from Aplysia giant (R2) cell somata showed long lasting 4-10 mV hyperpolarizations after passage of outward current through a second intracellular electrode.2. An increase in membrane slope conductance occurred simultaneously with the post-stimulus hyperpolarization (PSH).3. Both the PSH and conductance-increase varied strongly with stimulus amplitude and duration.4. Both the PSH and the conductance increase occurred in Ca-free medium containing tetrodotoxin, when action-potential production was completely blocked.5. The PSH persisted in the presence of ouabain or DNP, with cooling, with removal of external K(+), and in media where all the Na(+) was replaced with Li(+), suggesting that it was not due to the activity of an electrogenic pump.6. A reversal potential for the PSH was demonstrated by application of maintained inward current following the end of an outward-directed stimulus.7. The PSH reversal potential varied with [K](o), but not with [Cl](o) or [Na](o), suggesting that the PSH was mainly due to an increase in K conductance.8. The PSH and the conductance increase were reduced strongly when all the Na(+) was replaced with Tris, and only slightly when Na(+) was replaced with sucrose.