Two different mechanisms of calcium spike modulation by dopamine

Dopamine (10 to 50 microM) modulates in two different ways the duration of the Ca2+-dependent action potential recorded in the cell body of identified neurons of the snail Helix aspersa. In some neurons (cells E13 and F1) dopamine increases the amplitude of their Ca2+-dependent spike plateau by decreasing the S-current (Klein, M., J.S. Camardo, and E. R. Kandel (1982) Proc. Natl. Acad. Sci. U.S.A. 79: 5713-5717), a K+ current controlled by cyclic AMP. In another neuron (cell D2), dopamine decreases the Ca2+-dependent plateau of the somatic action potential by evoking a decrease in Ca2+-current resulting from a decrease in Ca2+ conductance. Both modulatory effects could be observed in the same single neuron in which dopamine induces decreases of both the Ca2+ conductance and cyclic AMP-dependent K+ conductance. Nevertheless, in these cells (such as cell F5) dopamine only evokes a decrease of the amplitude of the Ca2+ spike plateau. Since the modulation of the duration of the Ca2+ action potential recorded in the neuronal soma has been shown to constitute a good model of events taking place at synaptic endings, it is suggested that these modulatory mechanisms evoked by dopamine may be involved in processes of presynaptic facilitation and inhibition.     

Neuropeptide Y actions and the distribution of Ca2+-dependent Cl- conductance in rat dorsal root ganglion neurons

Neuropeptide Y (NPY) increases the excitability of 'small', nociceptive, dorsal root ganglion (DRG) neurons. This effect, which may contribute to the etiology of 'neuropathic' pain, has been attributed to attenuation of Ca2+-sensitive K+ conductance(s) (gK,Ca) following suppression of Ca2+ entry via N-type Ca2+ channels. A problem arises with this conclusion because rat DRG neurons normally contain high intracellular Cl- and some of them express a Ca2+-dependent Cl- conductance (gCl,Ca). In this study, we find that in rat DRG neurons increasing intracellular Cl- does not attenuate the effect of 1 microM NPY because gCl,Ca is not found in 'small' DRG cells and the peptide failed to affect the gCl,Ca found in 'large' cells. Thus, the presence of gCl,Ca in a subpopulation of 'large' DRG neurons does not alter the conclusion that excitatory effects of NPY result from attenuation of gK,Ca.     

Hyperpolarizing action of enkephalin on neurons in the dorsal motor nucleus of the vagus, in vitro

Intracellular recordings were made from neurons of the dorsomotor vagal nucleus (DMV) in slices of rat medulla oblongata. [D-Ala2, D-Leu5]-enkephalin (DADLE), applied by perfusion (0.01-3 microM) or droplets, dose-dependently hyperpolarized 85% of the DMV neurons tested. The hyperpolarization, associated with a decrease in membrane resistance, persisted after elimination of synaptic activity by perfusion with Ca2(+)-free/high-Mg2+ solution or with 1 microM TTX solution. The opioid antagonist, naloxone, reversibly inhibited DADLE-induced hyperpolarization. The hyperpolarization depended on extracellular K+ concentration and reversed at about -90 mV. DADLE also decreased Ca2(+)-dependent spike duration and after-hyperpolarization (AHP). DAGO (a selective mu-receptor agonist), but not DPLPE (a selective delta-receptor agonist), mimicked DADLE's effects on membrane potential, Ca2(+)-dependent spike duration, and AHP. It is concluded that DADLE, through postsynaptic mu-type opioid receptors, hyperpolarized DMV neurons by increasing K+ conductance, which may have an inhibitory effect on DMV output. DADLE-induced decrease of spike duration and AHP was also mediated by mu-receptors and could have additional effects on functions of the DMV neuron by virtue of reduction in Ca2+ entry.     

Subpopulations of GABAergic and non-GABAergic rat dorsal horn neurons express Ca2+-permeable AMPA receptors.

Subpopulations of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors that are either permeable or impermeable to Ca2+ are expressed on dorsal horn neurons in culture. While both mediate synaptic transmission, the Ca2+ -permeable AMPA receptors provide a Ca2+ signal that may result in a transient change in synaptic strength [Gu, J.G., Albuquerque, C., Lee, C.J. & MacDermott, A.B. (1996) Nature, 381, 793]. To appreciate the relevance of these receptors to dorsal horn physiology, we have investigated whether they show selective expression in identified subpopulations of dorsal horn neurons. Expression of Ca2+-permeable AMPA receptors was assayed using the kainate-induced cobalt loading technique first developed by Pruss et al. [Pruss, R.M., Akeson, R.L., Racke, M.M. & Wilburn, J.L. (1991) Neuron, 7, 509]. Subpopulations of dorsal horn neurons were identified using immunocytochemistry for gamma-aminobutyric acid (GABA), glycine, substance P receptor (NK1 receptor) and the Ca2+-binding proteins, calretinin and calbindin D28K. We demonstrate that, in dorsal horn neurons in culture, kainate-induced cobalt uptake is selectively mediated by Ca2+-permeable AMPA receptors, and that a majority of GABA and NK1 receptor-expressing neurons express Ca2+-permeable AMPA receptors. GABAergic dorsal horn neurons are important in local inhibition as well as in the regulation of transmitter release from primary afferent terminals. NK1 receptor-expressing dorsal horn neurons include many of the projection neurons in the nociceptive spino-thalamic pathway. Thus, we have identified two populations of dorsal horn neurons representing important components of dorsal horn function that express Ca2+-permeable AMPA receptors. Furthermore, we show that several subpopulations of putative excitatory interneurons defined by calretinin and calbindin expression do not express Ca2+-permeable AMPA receptors.     

Serotonin and cyclic GMP both induce an increase of the calcium current in the same identified molluscan neurons

Serotonin (5-HT) has previously been shown to evoke an increase in the duration of the Ca2+-dependent spike of molluscan neurons by decreasing the S current (Klein et al., 1982), a K+ current controlled by cAMP. However, in a group of identified ventral neurons of the snail Helix aspersa in which 5-HT (1-10 microM) also prolonged the duration of the Ca2+-dependent action potential, no 5-HT-induced depression of S current or of any other outward current was observed. Instead, 5-HT was found to evoke the prolongation of the somatic spike by inducing an increase in Ca2+ membrane conductance. This 5-HT-induced increase of Ca2+-current was mimicked neither by the intracellular injection of cAMP nor by the extracellular application of forskolin (20 microM). In contrast, it was mimicked by the intracellular injection of cGMP and by the extracellular application of 100 nM zaprinast, a cGMP-phosphodiesterase inhibitor. The extracellular application of phorbol ester TPA (100 nM), an activator of protein kinase C, was also found to increase the Ca2+ current in the identified snail ventral neurons, but this enhancing effect had a different time course from that induced by 5-HT. These results indicate that there is a second mechanism for prolonging the Ca2+ spike of molluscan neurons, consisting of an increase in Ca2+ current, in which cGMP may play a role as second messenger.     

Intracellular Calcium and Control of Burst Generation in Neurons of Guinea-Pig Neocortex in Vitro

Response properties of neurons in brain slices of guinea pig parietal neocortex were examined following intracellular injection of the Ca2+ chelators, EGTA and BAPTA. Although chelator injection did not cause any consistent change in passive membrane properties, it did induce 81% of neurons encountered at all sub-pial depths to become 'bursters', in that just-threshold depolarizing current pulses triggered all-or-none bursts of 2 - 5 fast action potentials. Transition to 'burstiness' was associated with disappearance of an AHP and appearance of a DAP. Although chelator caused a slight increase in steady-state firing rate, marked accommodation persisted. Extracellular Co2+ or Mn2+ had an effect on steady-state firing rate similar to that of the intracellular chelators; however, exposure to these Ca2+ channel blockers also caused steady state depolarization, increased resting input resistance and time constant, and profound spike broadening. This treatment never induced transition to 'burstiness'. Chelator-injected neurons ceased to generate bursts when Ca2+ was replaced by Mn2+ in the Ringer's solution. During exposure to 10-6 M TTX and 20 mM TEA, 50 - 200 msec Ca2+ spikes followed brief depolarizing pulses. As chelator was injected into the cell, there was progressive prolongation of the Ca2+ plateaus, which was associated with slowing of the rate at which membrane resistance gradually recovered following the initial increase in conductance. These findings indicate that under normal conditions, activity-related increases in intracellular Ca2+ activate processes which prevent most neocortical neurons from being bursters. These processes probably include Ca2+-dependent K+ currents, and Ca2+-dependent Ca2+ channel inactivation.     

Active and Passive Membrane Properties of Spinal Cord Neurons that Are Rhythmically Active during Swimming in Xenopus Embryos

Cellular properties have been examined in ventrally located Xenopus spinal cord neurons that are rhythmically active during fictive swimming and presumed to be motoneurons. Resting potentials and input resistances of such neurons are - 75 +/- 2 mV (mean +/- standard error) and 118 +/- 17 M ohm respectively. Most cells fire a single impulse, 0.5 to 2.0 ms in duration and 48.5 +/- 1.8 mV in amplitude, in response to a depolarizing current step. A minority fire several spikes of diminishing amplitude to more strongly depolarizing current. Cells held above spike, threshold fire on rebound from brief hyperpolarizing pulses. Spikes are blocked by 0.1 to 1.0 microM tetrodotoxin (TTX) and are therefore Na+-dependent. Current/voltage (I/V) plots to injected current are approximately linear near the resting potential but become non-linear at more depolarized levels. Cells recorded in TTX with CsCI-filled microelectrodes show a linearized I/V plot at depolarized membrane potentials suggesting the normal presence of a voltage-dependent K+ conductance activated at relatively depolarized levels. Most cells recorded in this way but without TTX fire long trains of spikes of near constant amplitude, pointing to a role of the K+ conductance in limiting firing in normal cells. Spike blockage with TTX reveals, in some cells, a transient depolarizing Cd2+-sensitive and therefore presumably Ca2+-dependent potential that increases in amplitude with depolarization. Cells in TTX, Cd2+, and strychnine, and recorded with CsCI-filled microelectrodes to block active conductances respond to hyperpolarizing current steps with a two component exponential response. The cell time constant (tau0) obtained from the longer of these by exponential peeling is relatively long (mean 15.7 ms). These findings contribute to an increased understanding of the cellular properties involved in spinal rhythm generation in this simple vertebrate.     

FMRF-amide-like substances in the leech. II. Bioactivity on the heartbeat system

In the preceding paper (Kuhlman, J. R., C. Li, and R. L. Calabrese (1985) J. Neurosci. 5: 2301-2309) FMRF-amide-like immunoreactivity was localized to a specific set of neurons in the leech. Three types of these neurons are involved in controlling the animal's heartbeat: HE motor neurons and HA modulatory neurons which directly innervate the hearts, and the swim-initiating interneurons (cells 204) which can accelerate the heartbeat central pattern generator. Application of synthetic FMRF-amide had effects on the hearts and the heartbeat central pattern generator that mimicked the actions of the HA and cell 204 neurons. Bath application of FMRF-amide (10(-7) to 10(-6) M) to the hearts activated their myogenic rhythm and increased their beat tension, thus mimicking the effects of activity in HA cells. Bath application of lower concentrations of FMRF-amide (10(-9) to 10(-8) M) to the isolated central nervous system dramatically accelerated the central motor program for heartbeat, thus mimicking the effects of activity in cell 204. These observations suggest that an FMRF-amide-like substance may be used as a chemical signal by HA and cell 204 neurons. The role of the FMRF-amide-like substance contained in HE motor neurons remains unclear, but it may be released along with the HE cell's neuromuscular transmitter, acetylcholine.     

Participation of calcium-dependent potassium conductance in the processes of membrane hyperpolarization of the pyramidal neurons in the sensorimotor cortex in the cat

In acute experiments on immobilized cats intracellular injection of Ca+ decreased of IPSP and postburst hyperpolarization amplitudes in pyramidal neurons of the sensorimotor cortex. Intracellular injection of ethylene glycol tetraacetic acid had almost the same effect. This substance also reduced the late part of spike afterhyperpolarization, while the early part remained practically unchanged. It is concluded that Ca2+-dependent K+-conductance might play an important role in the genesis of IPSP, postburst and spike afterhyperpolarization in the membrane of pyramidal neurons of the cat sensorimotor cortex.     

Prostaglandins block a Ca-dependent slow spike afterhyperpolarization independent of effects on Ca influx in visceral afferent neurons

The blockade of a slow Ca2+-activated K+-dependent afterhyperpolarization (AHPs) in rabbit visceral sensory neurons by the prostaglandins, PGE1 and PGD2, was investigated to determine whether the blockade was indirectly due to a reduction in Ca2+ influx. The prostaglandins (PGs) could block the AHPs in the absence of any change in Ca2+-dependent spikes elicited in the presence of tetrodotoxin and tetraethylammonium bromide. A PG-induced decrease in Ca2+-dependent spike width observed in some neurons was temporally dissociated from the PG-induced block of the AHPs. In addition, a slow afterhyperpolarization produced by the application of the Ca2+ ionophore, A23187, was blocked by the PGs. It is concluded that a reduction in Ca2+ influx is not responsible for the PG-induced blockade of the AHPs.     

Calcium-activated chloride conductance in frog olfactory cilia

We have measured the effects of cytoplasmic Ca2+ on the conductance of single cilia excised from frog olfactory receptor neurons. When free cytoplasmic Ca2+ is buffered at 0.1 microM, ciliary conductance is low. As Ca2+ is increased, ciliary conductance increases. Maximal conductance averages sevenfold higher than that measured in the absence of Ca2+. We estimate that the K1/2 for Ca2+ activation is 5 microM; the dose-response curve indicates some positive cooperativity of Ca2+ binding. Activation by Ca2+ is rapid and fully reversible. Most of the Ca(2+)-activated current is carried by Cl- and persists in the absence of Na+ and K+. The Cl- channel inhibitor 3',5-dichlorodiphenylamine-2-carboxylate (300 microM) reduces the Ca(2+)-activated current by 90%. Odorants induce a Ca2+ influx in some olfactory receptor neurons, but the consequences of this influx for neuronal function are not well understood. Our findings allow us to predict that a Ca2+ influx would increase the permeability of the olfactory cilia to Cl-. How this would affect the neuronal potential is uncertain, since the equilibrium potential for Cl- in olfactory receptor neurons is unknown.     

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 Ca2+, high Mg2+ media. Baclofen actions on LC neurons are resistant to the GABAA 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 Ba2+ to the external media. Taken together, these data suggest that baclofen directly hyperpolarizes LC neurons by activation of GABAB receptors which leads to an increase in K+ conductance.     

Actions of serotonin recorded intracellularly in rat dorsal lateral septal neurons

The actions of serotonin (5HT) on passive and active membrane properties of neurons in the rat dorsal lateral septal nucleus (LSN) were studied by using intracellular recordings in transverse, septal slices. Superfusion with 10 microM 5HT induced a hyperpolarization of the membrane in almost all neurons tested in the dorsolateral part of the LSN. The hyperpolarization was accompanied by a decrease in membrane resistance. These effects of 5HT persisted in a low-Ca2+/high-Mg2+-containing medium or medium with tetrodotoxin, indicating a post-synaptic site of action for 5HT. The reversal potential for the hyperpolarizing effect was ca. -95 mV. If the extracellular K+-concentration was raised, the reversal potential became less negative. These data suggest that 5HT hyperpolarizes LSN neurons by increasing a K+-conductance. Spontaneous, synaptically evoked action potentials and action potentials induced in LSN neurons by a depolarizing current step typically display a fast Na+-spike with a subsequent K+-afterhyperpolarization, followed by a much slower Ca2+-dependent afterdepolarization. The amplitude of the K+-afterhyperpolarization was decreased by 5HT, while at the same time the afterdepolarization became more pronounced. The Ca2+-spike of LSN neurons was not affected by 5HT. Synaptic responses that were evoked in LSN neurons by stimulation of the dorsal part of the LSN consisted of a fast EPSP or spike, followed by a Cl(-)-dependent fast IPSP and a K+-dependent late IPSP. Of these synaptic responses, 5HT suppressed particularly the late IPSP. The present data indicate that 5HT affects the conductance for active and passive K+-channels in LSN neurons.(ABSTRACT TRUNCATED AT 250 WORDS)     

The beta-amyloid precursor protein controls a store-operated Ca2+ entry in cortical neurons

A polyclonal antibody (APP-Ab) raised against the extracellular domain of the beta-amyloid precursor protein (APP) triggers a marked neuronal cell death preceded by activation of Ca(2+)-dependent enzymes, neurite degeneration, oxidative stress and nuclear condensation [Mbebi et al. (2002) J. Biol. Chem., 277, 20979-20990]. We have investigated whether activation of APP by this antibody could promote cell death through cellular Ca2+ homeostasis alteration. We carried out time-lapse recordings of intracellular Ca2+ signals in cultured mice cortical neurons by means of a scanning confocal microscope. When applied in the presence of 2 mm external Ca2+, APP-Ab elicited a long-lasting elevation of the intracellular concentration of Ca2+ ([Ca2+]i). Experiments performed in the absence of external Ca2+ showed that APP-Ab triggers the release of Ca2+ from intracellular stores. The re-admission of external Ca2+ provides an additional rise of Ca2+ most likely through store-operated channels. A pretreatment of the cells with pertussis toxin, to inhibit the activity of Gi/Go proteins, or with the phospholipase C inhibitor, 3-nitrocoumarin, prevented both the APP-dependent elevation of Ca2+ as well as the APP-Ab-mediated cell death. Similarly, the store-operated channel inhibitors, 2-APB or SKF-96365 block both the APP-Ab-dependent Ca2+ entry and the APP-Ab-mediated cell death. Altogether, our data provide functional evidence that APP can perturb intracellular Ca2+ homeostasis by emptying intracellular Ca2+ stores and triggering Ca2+ entry through store-operated channels. In response to APP activation, the long-lasting elevation of [Ca2+]i due to an entry of Ca2+ via store-operated channels appears as a major event that leads to neuronal cell death.     

Low-dose benzodiazepine neuronal inhibition: enhanced Ca2+-mediated K+-conductance

The water-soluble inhibitory benzodiazepine, midazolam, was applied in low nanomolar concentrations to CA1 hippocampal neurons in vitro, recorded intracellularly. The drug caused a long-lasting hyperpolarization and moderate conductance increase, which persisted with TTX-induced synaptic blockade or with intracellular injection of Cl- ions, but not in zero Ca2+ perfusate. Calcium spikes elicited in the presence of TTX were enhanced by midazolam. It was concluded that these low nanomolar concentrations, which did not enhance GABA actions, inhibited by augmenting Ca2+ mediated K+-conductance.     

Activation and modulation of neuronal K+ channels by GABA.

Although the concept of GABAB receptors was introduced only ten years ago, several actions of GABAB agonists are already well established. They cause depression of transmitter release, a decrease in voltage-dependent Ca2+ conductance and an increase in K+ conductance. It has recently been reported that GABA also changes the voltage dependence of the transient ('A' type) K+ channel. Depression of transmitter release by GABAB agonists may be caused by a decrease in Ca2+ conductance, an increase in K+ conductance or a modulation of A channels in presynaptic nerve terminals. Slow IPSPs in some neurons are generated by an increase in K+ conductance that can be blocked by GABAB antagonists and pertussis toxin. K+ channels of variable amplitude that are blocked by pertussis toxin are activated by GABAB agonists in cultured hippocampal neurons. Since arachidonic acid activates similar channels in excised patches of membrane, it may form part of a normal second messenger system linking GABAB receptors to K+ channels.     

The large conductance Ca2+-activated potassium channel (pSlo) of the cockroach Periplaneta americana: structure,localization in neurons and electrophysiology

Voltage-activated, Ca2+-sensitive K+ channels (BK or maxi K,Ca channels) play a major role in the control of neuronal excitability. We have cloned pSlo, the BK channel alpha subunit of the cockroach Periplaneta americana. The amino acid sequence of pSlo shows 88% identity to dSlo from Drosophila. There are five alternatively spliced positions in pSlo showing differential expression in various tissues. A pSlo-specific antibody prominently stained the octopaminergic dorsal unpaired median (DUM) neurons and peptidergic midline neurons in Periplaneta abdominal ganglia. HEK293 cells expressing pSlo exhibit K+ channels of 170 pS conductance. They have a tendency for brief closures, exhibit subconductance states and show slight inward rectification. Activation kinetics and voltage dependence are controlled by cytoplasmic [Ca2+]. In contrast to dSlo, pSlo channels are sensitive to charybdotoxin and iberiotoxin. Mutagenesis at two positions (E254 and Q285) changed blocking efficacy of charybdotoxin. In contrast to pSlo expressed in HEK293 cells, native IbTx-sensitive K,Ca currents in DUM and in peptidergic neurons, exhibited rapid, partial inactivation. The fast component of the K,Ca current partly accounts for the repolarization and the early after-hyperpolarization of the action potential. By means of Ca2+-induced repolarization, BK channels may reduce the risk of Ca2+ overload in cockroach neurons. Interestingly, the neurons expressing pSlo were also found to express taurine, a messenger that is likely to limit overexcitation by an autocrine mechanism in mammalian central neurons.     

Soman reversibly decreases the duration of Ca and Ba action potentials in bullfrog sympathetic neurons

Soman, (pinacoloxymethyl-phosphoryl fluoride) (0.1–10 μM) an irreversible cholinesterase inhibitor, reversibly reduced the duration of calcium (Ca²⁺)- and barium (Ba²⁺) spikes without significantly affecting spike amplitude in sympathetic postganglionic neurons of the adult bullfrog (Rana catesbeiana). The soman-induced shortening of the spike duration was not prevented by pretreatment with either (+)-tubocurarine (100 μM) or hexamethonium (100 μM) and atropine (10 μM) and was also recorded from acutely-dissociated sympathetic neurons. These results suggest that soman has a direct action to decrease calcium entry through voltage-dependent channels activated during a spike. This effect may contribute to both the decrease in the duration of the spike after-hyperpolarization (AHP) and the enhanced neuronal excitability produced by soman in these neurons.     

Electrical membrane properties of rat substantia nigra compacta neurons in an in vitro slice preparation

The electrical membrane properties of rat substantia nigra pars compacta (SNC) neurons were studied in an in vitro slice preparation. Some of the recorded neurons were intracellularly labeled with HRP and were found to have morphological characteristics resembling the presumed SNC dopaminergic neurons, as reported by others. The input resistance of SNC neurons at resting membrane potential ranged between 70 and 250 M omega. The membrane resistance showed strong anomalous rectification when the membrane was hyperpolarized by current injection. The anomalous rectification was decreased by the addition of tetraethylammonium bromide (TEA) to the bathing Ringer solution. Injection of depolarizing current or termination of hyperpolarizing current induced slow depolarizing potentials. Their amplitude was dependent on the membrane potential and the current intensity. In neurons treated with tetrodotoxin (TTX) and TEA, slow action potentials were triggered from the slow depolarizing potentials. Both the slow depolarizing potential and slow action potential were TTX resistant and abolished by superfusion of Ca2+-free medium. Long duration hyperpolarizations were observed following the injection of depolarizing current pulses. The hyperpolarization was abolished by the superfusion of Ca2+-free medium or decreased by addition of TEA to the Ringer solution indicating an involvement of a Ca2+-dependent K+-conductance in generation of the hyperpolarization. The long duration hyperpolarization was also observed following action potentials. The spike after hyperpolarization consisted of an initial short duration fast component and a long lasting component. The amplitude of both components seems to be reduced but not abolished by TEA (up to 10 mM). When hyperpolarizing current pulses were applied to neurons that were held either continuously depolarized or were superfused with Ca2+-free medium, the pattern of the membrane potential after the offset of current pulses consisted of an initial fast and a later slow ramp-shaped phase. The latter was associated with a membrane conductance increase and interpreted to be due to an early K+ current. This early K+ current was relatively resistant to TEA. Injections of strong depolarizing currents triggered action potentials with multiple inflections on their rising phase. The amplitudes of action potentials changed abruptly during current application. These data indicate that SNC neurons have multiple generation sites for action potential.     

Intracellular injection of apamin reduces a slow potassium current mediating afterhyperpolarizations and IPSPs in neocortical neurons of cats

Electrophysiologic effects of intracellularly injected apamin, a Ca2+-dependent K+ channel blocker, were investigated in neurons of the motor cortex of awake cats. Single-electrode voltage clamp techniques were used to measure changes in membrane currents including those that were synaptically activated. All changes occurred within 2-4 min after pressure injection of apamin with partial recovery observed within 8-15 min. Apamin selectively abolished an outward current that mediated a slow afterhyperpolarization (AHP) following intracellular depolarizing current pulses and action potentials without influencing the time course of the action potentials or an associated fast AHP component. In addition apamin increased the number and frequency of spike discharges evoked by the depolarizing current pulses and produced a small increase in the rate of background firing activity. The baseline resting potential and input resistance were essentially unchanged by apamin. Apamin also diminished a late, slowly decaying component of inhibitory postsynaptic potentials (IPSPs) and currents (IPSCs) elicited by stimulation of the ventrolateral thalamus or the pyramidal tract. The apamin-induced changes were concomitant with a decrease of the decay time constant of both IPSPs and IPSCs and a positive shift in their reversal potential. The results suggest that the late, slowly decaying component of these inhibitory postsynaptic responses is generated by an apamin-sensitive Ca2+-dependent K+ conductance which is also responsible for the slow AHP.