A ouabain-sensitive hyperpolarization in rat striatal neurones in vitro

Intracellular recordings were made from rat striatal neurones in vitro. In the presence of intracellular caesium and extracellular tetraethylammonium chloride (TEA) (5 mM) and barium (3 mM), long-lasting plateau potentials developed followed by a prominent voltage independent hyperpolarization which lasted several seconds. A similar afterhyperpolarization was observed when calcium was replaced by barium. The afterhyperpolarization was reduced in a potassium free medium and reversibly abolished in a Na+-free solution or by cooling the slice to 21-24 degrees C. It was also irreversibly blocked by ouabain (50 microM). This hyperpolarization may therefore result from the activation of a Na+,K+-ATPase electrogenic pump.     

Excessive intracellular Ca2+ inhibits glutamate-induced Na(+)-K+ pump activation in rat hippocampal neurons.

1. The effects of increased intracellular Ca2+ concentration ([Ca2+]i) on Na(+)-K+ pump activity in CA1 pyramidal neurons of rat hippocampal slices were investigated. The postglutamate hyperpolarization (PGH), which follows glutamate (GLU)-induced depolarization (GD), was used as an index of Na(+)-K+ pump activity, as was a ratio of PGH area to the preceding GD area (PGH ratio). 2. Perfusion of slices with saline containing Ca2+ ionophore (A23187, 10 microM) inhibited the PGH without producing apparent signs of cell deterioration. A 60-100% (85 +/- 15%, mean +/- SD) reduction in the PGH ratio occurred after 20-50 min of A23187 superfusion in 12 of 18 neurons tested. Complete abolition of the PGH occurred in 8 of these 12 cells exposed to A23187 for 30-120 min. 3. Application of A23187 in Ca(2+)-free/high-Mg2+ solution did not abolish the PGH, although small (less than 50%; 37 +/- 10%) reductions in the PGH ratio were observed after perfusion of 50 min or longer in five neurons tested. 4. Intracellular injection of the Ca2+ chelator bis-(o-amino-phenoxy)-N,N,N',N'-tetraacetic acid (BAPTA, 300-400 mM) blocked inhibition of the PGH by A23187. After 50 min of perfusion with Ca2+ ionophore, no reduction of the PGH ratio was observed in five neurons tested. 5. Rundown of the PGH without apparent change in membrane properties was observed in three neurons that were stable for greater than 2-3 h, allowing repetitive GLU applications. 6. Block of the PGH produced by a Na(+)-K(+)-adenosinetriphosphatase (ATPase) inhibitor (strophanthidin) prolonged the duration of GDs because of a delay in repolarization.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conductance changes, an electrogenic pump and the hyperpolarization of leech neurones following impulses

Following trains of impulses, sensory neurones in the C.N.S. of the leech show a prolonged hyperpolarization, which lasts for seconds or minutes. In the present investigation the mechanisms that underly this hyperpolarization have been studied by recording intracellularly. Two factors have been found to be responsible. One is the activity of an electrogenic pump (see Baylor & Nicholls, 1969b); the other is a long-lasting change in K conductance.1. Additional evidence that an electrogenic pump contributes to a slow after-hyperpolarization of leech sensory neurones is provided by the effects of injecting Na intracellularly. This leads to an increase in membrane potential that is blocked by the cardiac glycoside strophanthidin. Furthermore, after a train of impulses, reducing the K concentration in the external fluid characteristically reduces the hyperpolarizing action of the pump.2. The hyperpolarization following impulses is associated with a reduction of the cell membrane resistance that can persist for several minutes.3. Several lines of evidence suggest that the reduction in input resistance during the hyperpolarization is mainly due to an increased permeability to K. Thus, when the K concentration in Ringer fluid is reduced, the peak amplitude of the hyperpolarization following a train becomes larger. Furthermore, the conductance dependent part of the after-hyperpolarization has a reversal potential close to the equilibrium potential for K (E(K)). Substitution of Cl by SO(4) has little effect either on the after-hyperpolarization or on the conductance change following a train.4. Increased external Ca concentrations lead to a marked increase in the hyperpolarization that follows impulse activity. The enhanced hyperpolarization in high Ca is associated with a corresponding reduction in input resistance. The amplitude and duration of the hyperpolarization following a brief train of impulses can be increased by a factor of 5 or more in Ringer fluid containing 10 mM-Ca instead of the usual 1.8 mM. The hyperpolarization and resistance changes still occur in solutions containing 20 mM-Mg.5. To augment the hyperpolarization the increased concentration of Ca must be present during the train of impulses.6. The relative contributions of the K conductance increase and of the electrogenic pump for generating the hyperpolarization after impulse activity are different in the three types of sensory cell responding to touch, pressure and noxious stimulation.     

Post-tetanic hyperpolarization produced by an electrogenic pump in dorsal spinocerebellar tract neurones of the cat

1. Hyperpolarization following single and repetitive excitation of dorsal spinocerebellar tract (DSCT) neurones of the cat was studied by intracellular recording.2. Hyperpolarization following an antidromic action potential consisted of an initial, brief phase (undershoot) and a late, prolonged phase. The latter hyperpolarization was independent of the membrane potential, whereas the former was reversed in polarity by hyperpolarizing pulses applied across the DSCT cell membrane.3. DSCT neurones showed a prolonged hyperpolarization after a train of antidromic action potentials. The amplitude and duration of the hyperpolarization were dependent on the number and the frequency of action potentials. A similar hyperpolarization was observed following a train of impulses evoked by depolarizing pulses applied through the intracellular electrode.4. There was no detectable conductance change during the post-tetanic hyperpolarization. The latter showed no reversal potential when the membrane potential was altered.5. The half-decay time of the post-tetanic hyperpolarization was lengthened when the cord temperature was lowered. The temperature coefficient (Q(10)) was 2.4 within the range of 31-40 degrees C.6. The amplitude of the undershoot following each action potential was assumed to provide a criterion for the accumulation of the extracellular K(+). Alterations in the amplitude of undershoots during repetitive excitation suggested that the duration of post-tetanic hyperpolarization depends on the accumulation of extracellular K(+) as well as of intracellular Na(+) associated with a train of impulses.7. It is suggested that post-tetanic hyperpolarization is produced by an electrogenic sodium pump. A possible significance of such a hyperpolarization in impulse coding is discussed.     

Contribution of an electrogenic sodium pump to the membrane potential in rabbit sinoatrial node cells

A study has been made of the transient hyperpolarization (K+-induced hyperpolarization) which developed following readmission of potassium after having pre-treated the rabbit sinoatrial node tissue with K+-depleted Tyrode solution for 4--5 min at 35 degrees C. Evidence is presented indicating that the K+-induced hyperpolarization results from the activity of an electrogenic sodium pump: The K+-induced hyperpolarization was inhibited by substituting Li+ for Na+ and by cooling the tissue. The amplitude of the K+-induced hyperpolarization was increased either by increasing K+ concentration in the recovery solution or by decreasing K+ concentration in the pre-treatment K+-depleted solution. By removing Cl- from the perfusates, the amplitude of the K+-induced hyperpolarization increased. In a Cl--depleted solution, the sinoatrial node cell membrane hyperpolarized by approximately 15 mV without a transient depolarization.     

Electrogenic pump (Na+/K(+)-ATPase) activity in rat optic nerve

Rat optic nerves were studied in a sucrose gap chamber in order to study the origin of a late afterhyperpolarization that follows repetitive activity. The results provide evidence for electrogenic pump (Na+/K(+)-ATPase) activity in central nervous system myelinated axons and demonstrate an effect on axonal excitability. Repetitive stimulation (25-200 Hz; 200-5000 ms) led to a prolonged, temperature-dependent post-train afterhyperpolarization with duration up to about 40 s. The post-train afterhyperpolarization was blocked by the Na+/K(+)-ATPase blockers strophanthidin and ouabain, and the substitution of Li+ for Na+ in the test solution, which also blocks Na+/K(+)-ATPase. The peak amplitude of the post-train afterhyperpolarization was minimally changed by the potassium-channel blocker tetraethylammonium (10 mM), and the Ca2(+)-channel blocker CoCl2 (4 mM). Hyperpolarizing constant current did not reverse the afterhyperpolarization. The amplitude of the hyperpolarization was increased in the presence of the potassium-channel blocker 4-aminopyridine (1 mM). In the presence of 4-amino-pyridine, the post-train hyperpolarization was much reduced by strophanthidin, except for a residual early component lasting several hundred milliseconds which was blocked by the potassium-channel blocker tetraethylammonium. This finding indicates that after exposure to 4-aminopyridine, repetitive stimulation leads to activation of a tetraethylammonium-sensitive K(+)-channel that contributes during the first several hundred milliseconds to the post-train afterhyperpolarization. The amplitude of the compound action potential elicited by a single submaximal stimulus during the post-train hyperpolarization was smaller than that of the control response.The decrement in amplitude was not present under identical stimulation conditions when the post-train hyperpolarization was blocked by strophanthidin, indicating that the hyperpolarization associated with repetitive stimulation reduced excitability.(ABSTRACT TRUNCATED AT 250 WORDS)     

Adrenaline-induced K+ efflux results in sodium pump stimulation in a sympathetic ganglion

The potassium-activated hyperpolarization (KH) was used as an index of electrogenic Na+ pumping in bullfrog sympathetic ganglia. This response was evoked by storing ganglia in K-free Ringer's solution and briefly introducing normal Ringer's solution containing 2 mM K+ at regular intervals. The apparent EC50 for K+ was 2.21 mM (range 0.88-3.54 mM, for n = 5) and at least 10 mM K+ was required to produce a maximal KH response. Adrenaline, which produces membrane hyperpolarization by increasing K+ conductance (gK), increased the amplitude of KH responses. When the K+ efflux accompanying the adrenaline-induced hyperpolarization (AdH) was blocked with 2 mM Ba2+, the KH was no longer potentiated. It is suggested that the K+ moving out of the cells during the AdH accumulates extracellularly and stimulates the Na+ pump.     

Cyclic variation of potassium conductance in a burst-generating neurone in Aplysia

1. The hyperpolarization between bursts in the R 15 cell of Aplysia is accompanied by an increase in membrane slope conductance.2. The post-burst hyperpolarization can be observed with ouabain, lithium, or potassium-free solution if artificial inward current is applied. The hyperpolarization can be observed with dinitrophenol or cooling to 10 degrees C, with no injected current. Thus, the hyperpolarization apparently is not due to the cyclic activity of an electrogenic pump.3. A reversal potential for the post-burst hyperpolarization can be demonstrated by passage of inward current during the inter-burst period. The reversal of direction of the potential depends on recent occurrence of a burst.4. The reversal potential varies with external potassium concentration, but not with chloride or sodium.5. The post-burst hyperpolarization is not blocked by external tetraethylammonium at a concentration which greatly prolongs the action potentials.6. During the onset of spike blockage by, and recovery from, calcium-free+tetrodotoxin saline, the bursts of action potentials appear to be driven by endogenous waves of membrane potential.7. The hyperpolarizing phase of the waves in calcium-free+tetrodotoxin medium is accompanied by an increased slope conductance.8. A reversal potential can be demonstrated for the hyperpolarization following a wave in calcium-free+tetrodotoxin medium by applying inward current during the interwave period.9. The waves in calcium-free+tetrodotoxin medium are blocked by ouabain but can be reinstated by artificial hyperpolarization.10. The post-burst hyperpolarization and the post-wave hyperpolarization appear to result from a periodic increase in membrane conductance, primarily to potassium ions.     

Inhibitory action of serotonin in CA1 hippocampal neurons in vitro

The ionic mechanism of the inhibitory effect of serotonin was investigated in vitro in the CA1 region of the rat hippocampus by extra- and intracellular recordings. Local or bath applications of serotonin induced a long-lasting reduction of extracellularly recorded synaptic potentials and orthodromic population spikes without affecting the afferent volley or the antidromic population spike. Serotonin can also reduce the frequency of occurrence of spontaneous excitatory and inhibitory postsynaptic potentials without any reduction of input resistance of the pyramidal neuron. During the response to serotonin, the conductance increase evoked by GABA, the inhibitory neurotransmitter, was not changed. A direct postsynaptic effect of serotonin was demonstrated: local or bath applications of serotonin induced a tetrodotoxin-resistant hyperpolarization and conductance increase. The conductance change was not reduced by manual clamp of the neurons to the control resting membrane potential; therefore, a possible involvement of the sodium-potassium electrogenic pump is unlikely. When neurons were loaded with chloride, serotonin could still induce a hyperpolarization with an apparent reversal more negative than the resting membrane potential. When neurons were loaded with caesium, the hyperpolarization and the conductance increase evoked by serotonin were blocked. It is therefore concluded that serotonin increases potassium permeability. Similar effects were induced by a 5-HT1A ligand. The slow after hyperpolarization was reduced by serotonin; the calcium spike was reduced at the same time. In caesium loaded neurons, the spike duration was not modified by serotonin. In the presence of extracellular caesium (4-5 mM), the serotonin-induced hyperpolarization and the conductance change were blocked, but the effect of serotonin on calcium spikes persisted. Tetraethylammonium (5-10 mM) or 4-aminopyridine (0.5 mM) had no effect on the response to serotonin. These data indicate that serotonin has a postsynaptic inhibitory action by an activating potassium conductance. The possibility of a regulation of calcium currents is discussed. The possible role of serotonin on intrinsic synaptic transmission is also discussed.     

Evidence for bursting pacemaker neurones in cultured spinal cord cells

Intracellular recordings were made from dissociated mouse spinal cord cells in primary culture. One type of spinal cord neurone, with a large cell body (40-50 micron), 3-5 short neurites, and a mean resting potential of -65 mV, was found to fire rhythmic bursts of action potentials with a phase duration of approximately 1s when the membrane potential was depolarized to -55 mV. These bursts did not arise from spontaneous synaptic input, but appeared to result from endogenous ionic conductance properties of the membrane resembling those observed in molluscan bursting pacemaker neurones. Ionic conductances underlying this bursting activity were studied pharmacologically by local application of ionic conductance blockers. Pacemaker potentials depended on Na+ conductance, since tetrodotoxin and Na-free medium were the most potent agents for blocking spontaneous rhythmic activity. However, a Ca2+ conductance was involved in the depolarizing phase of membrane potential oscillations, since Ba2+ application increased oscillation amplitude. Action potentials observed during the bursts were Na+- and Ca2+-dependent. They did not differ significantly from those observed in other spinal cord neurones in culture. Application of tetraethylammonium, CoCl2, BaCl2 and 4-aminopyridine revealed at least three different potassium conductances which controlled this bursting pacemaker activity. A delayed potassium conductance controlled spike duration, a Ca-dependent potassium conductance controlled the duration of the burst and underlay the hyperpolarizing phase terminating the burst, and finally, a transient potassium conductance appeared to be involved in the control of phase duration. The demonstration that spinal cord neurones growing in monolayer culture display typical bursting pacemaker activity raises the possibility that bursting pacemaker neurones in the mammalian spinal cord may be involved in a phasic pattern generator that could control such activities as walking and the respiratory rhythm.     

Calcium-dependent hyperpolarizations in bullfrog sympathetic neurons

Intracellular recordings were made from neurons in bullfrog sympathetic ganglia. Fast B and slow B neurons were identified and selected for the analysis [Dodd and Horn (1983) J. Physiol., Lond. 334, 255-269]. A single soma action potential was followed by a prolonged afterhyperpolarization lasting for several hundred ms up to 2 s. Part of the spike afterhyperpolarization was due to potassium conductance activation triggered by calcium entry during an action potential. Acetylcholine was directly applied onto the soma membrane by iontophoresis. A rapid nicotinic depolarization was followed by a slow muscarinic depolarization. The nicotinic depolarization was followed by a hyperpolarization when the muscarinic depolarization was blocked by scopolamine. This hyperpolarization was several mV in amplitude and from 1 to 10 s in duration. It disappeared when the preceding nicotinic depolarization was blocked by (+)-tubocurarine. A single fast excitatory postsynaptic potential was also followed by a hyperpolarization in the presence of scopolamine. The acetylcholine-induced hyperpolarization was due to potassium conductance activation triggered by calcium entry during the nicotinic depolarization. The present findings show that non-synaptic autoinhibition is operating in sympathetic neurons. In other words, a rapid nicotinic transmission leads to prolonged hyperpolarizations which are not mediated by any transmitters but are mediated by calcium.     

Pancreatic islet cells: electrogenic and electrodiffusional control of membrane potential.

1. Responses of the membrane electrical characteristics of mouse pancreatic islet cells to ionic environmental changes have been used to assess the role of [Na]0 and [K]0 in the control of membrane potential, i.e. by electrodiffusion or via an electrogenic sodium pump. Islet cell electrical properties were measured in vitro with intracellular glass micro-electrodes. 2. Substitution of LiCl for extracellular NaCl did not change the islet cell membrane potential significantly in low (2.8 mM) glucose solutions, but readmission of NaCl caused a transient hyperpolarization (membrane potential maximum: -70 mV) in high glucose; when choline chloride was substituted for NaCl no hyperpolarization was observed on NaCl re-admission. 3. Superfusion with K-free solution gave no marked change in membrane potential during 30 min incubation in either low (2-8 mM) or high (28 mM) glucose concentrations but longer periods of exposure to K-free solutions caused progressive depolarization. 4. Readmission of K+ induced a transient hyperpolarization of up to 30 mV magnitude and 10 min duration in the presence of high (28 mM) but not low glucose (2-8 mM) concentrations. At the level of maximum hyperpolarization the membrane potential reached -60 mV, the electrical activity induced by the high glucose concentration being concurrently completely blocked. Replacement of [Cl]0 by isethionate accentuated these effects. 5. Ouabain, 10(-3) M, or a decrease in temperature from 37 to 7 degrees C depolarized the islet cells and blocked the transient hyperpolarization induced by readmission of K+. 6. Diphenylhydantoin, 1-5 times 10(-4) M, caused a significant hyperpolarization of the islet cells in low glucose (2-8 mM) and inhibited the electrical activity induced by high glucose (28 mM) or tolbutamide (0-7 mM). 7. It is concluded from these results that both an electrogenic and ionic component contribute to the membrane potential of the mouse pancreatic islet cell although electrodiffusional control normally predominates; acceleration of the Na-K exchange pump by diphenylhydantoin inhibits glucose-induced electrical activity. These findings are discussed in relation to the permeability characteristics of the islet cell membrane and the mechanism of insulin release.     

Dopamine modulates CA1 hippocampal neurons by elevating the threshold for spike generation: an in vitro study

Intracellular recordings were obtained from CA1 neurons of rat hippocampal slices preparation. Dopamine applied by perfusion (10(-5)-10(-7) M), microdrop (10(-4) M) and iontophoresis (+80, +200 nA balanced current) inhibited "spontaneous" and evoked action potentials. An increase in current injection restored the evoked action potentials which appeared unmodified. Membrane potential was not modified in 60% of the neurons; in the remaining ones, a slow depolarization was observed. Membrane resistance, measured at rest, was not modified by dopamine. Calcium-mediated events such as bursting activity and afterhyperpolarization, mainly in the late component, were also attenuated by the catecholamine. These effects were antagonized by domperidone, a dopaminergic antagonist. Calcium spikes, evoked in tetrodotoxin- and tetraethylammonium-poisoned slices, were reversibly inhibited by dopamine. Since an increase in the amplitude of a depolarizing pulse of injected current was able to evoke both sodium and calcium action potentials suppressed by dopamine without change in shape or duration, it is concluded that this catecholamine depresses cellular excitability by altering the interaction between membrane voltage and sodium and calcium entry and the subsequent increase in potassium conductance.     

Excitatory transmission in the basolateral amygdala.

1. Intracellular current-clamp recordings obtained from neurons of the basolateral nucleus of the amygdala (BLA) were used to characterize postsynaptic potentials elicited through stimulation of the stria terminalis (ST) or the lateral amygdala (LA). The contribution of glutamatergic receptor subtypes to excitatory postsynaptic potentials (EPSPs) were analyzed by the use of the non N-methyl-D-aspartate (non-NMDA) antagonist, 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX), and the NMDA antagonist, (DL)-2-amino-5-phosphonovaleric acid (APV). 2. Basic membrane properties of BLA neurons determined from membrane responses to transient current injection showed that at the mean resting membrane potential (RMP; -67.2 mV) the input resistance (RN) and time constant for membrane charging (tau) were near maximal, and that both values were reduced with membrane hyperpolarization, suggesting an intrinsic regulation of synaptic efficacy. 3. Responses to stimulation of the ST or LA consisted of an EPSP followed by either a fast inhibitory postsynaptic potential (f-IPSP) only, or by a fast- and subsequent slow-IPSP (s-IPSP). The EPSP was graded in nature, increasing in amplitude with increased stimulus intensity, and with membrane hyperpolarization after DC current injection. Spontaneous EPSPs were also observed either as discrete events or as EPSP/IPSP waveforms. 4. In physiological Mg2+ concentrations (1.2 mM), at the mean RMP, the EPSP consisted of dual, fast and slow, glutamatergic components. The fast-EPSP (f-EPSP) possessed characteristics of kainate/quisqualate receptor activation, namely, the EPSP increased in amplitude with membrane hyperpolarization, was insensitive to the NMDA receptor antagonist, APV (50 microM), and was blocked by the non-NMDA receptor antagonist, CNQX (10 microM). In contrast, the slow-EPSP (s-EPSP) decreased in amplitude with membrane hyperpolarization, was insensitive to CNQX (10 microM), and was blocked by APV (50 microM), indicating mediation by NMDA receptor activation. 5. In the presence of CNQX (10 microM), ST stimulation evoked an APV-sensitive s-EPSP. In contrast, LA stimulation evoked a f-IPSP, which when blocked by subsequent addition of bicuculline methiodide (BMI; 30 microM) revealed a temporally overlapping APV-sensitive s-EPSP. These data suggest that EPSP amplitude and duration are determined, in part, by the shunting of membrane conductance caused by a concomitant IPSP. 6. Superfusion of either CNQX or APV in BLA neurons caused membrane hyperpolarization and blockade of spontaneous EPSPs and IPSPs, suggesting that these compounds may act to block tonic excitatory amino acid (EAA) release within the nucleus, and that a degree of feed-forward inhibition occurs within the nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)     

A voltage- and time-dependent rectification in rat dorsal spinal root axons.

1. Rat dorsal spinal roots were studied by the use of whole-nerve sucrose gap and intra-axonal recording techniques. A prominent time-dependent conductance increase as evidenced by a relaxation or "sag" in membrane potential toward resting potential was elicited in dorsal spinal roots by constant hyperpolarizing current pulses. The relaxation, or sag, indicative of inward rectification, reached a maximal level and then decayed during the current pulse. 2. The time-dependent sag elicited by hyperpolarization was reduced when Na+ or K+ was removed from the normal bath solution but was abolished with the removal of both Na+ and K+. Tetrodotoxin (TTX), tetraethylammonium (TEA), and 4-aminopyridine (4-AP) did not affect the depolarization sag, suggesting that conventional voltage-dependent sodium and potassium channels do not underlie the inward rectification. 3. Cs+ in low concentrations completely abolished the inward rectification, whereas Ba2+ induced a partial block. 4. Current-voltage curves indicate that the magnitude of the depolarizing sag increases monotonically with increasing hyperpolarization. The time required to reach peak hyperpolarization, maximal sag potential, and the time between peak hyperpolarization and sag membrane potentials decreases with increasing levels of hyperpolarization. 5. The inward rectification is refractory to further stimulation during its decay phase, as revealed by paired-pulse protocols. This decay in inward rectification is both time and voltage dependent and is observed on a single axon level by the use of intra-axonal recording techniques as well as from whole-root recordings in the sucrose gap. 6. It is concluded that rat dorsal root fibers display a prominent time-dependent conductance increase in response to hyperpolarization that depends on both Na+ and K+ permeability and is blocked by Cs+. This rectification displays a decay phase that has not been previously described for similar conductances. It is argued that the Na+ component of this conductance is primarily responsible for stabilizing membrane potential near resting potential during periods of hyperpolarization.     

Ionic mechanisms of intestinal electrical control activity.

The effects of inhibition and stimulation of the electrogenic Na pump and of altering the ionic environment on the electrical control activity (ECA) were studied in rabbit jejunal smooth muscle. Pump inhibition abolished the ECA at a time when the membrane potential was more negative than the peak depolarization of the control potential (CP). Pump stimulation hyperpolarized the membrane and CP's appeared. Their amplitude was initially small and progressively increased as the hyperpolarization subsided. Lowering external Na to 20 mM or Ca withdrawal, but not addition of verapamil, reversibly abolished the ECA. Chloride replacement by propionate, isethionate, or benzene-sulphonate caused a transient augmentation, followed by suppression of the secondary depolarization of the CP's and decreased their frequency. The initial depolarization of the CP was little affected. Nitrate substitution increased CP frequency and spiking activity but had no observable effects on the CP configuration. These results suggest that the intestinal control potential may result from conductance changes initially to Na and later to C1 rather than fron an oscillatory electrogenic pump.     

The effect of caffeine on the neurons of a mammalian sympathetic ganglion.

The effect of caffeine on the isolated superior cervical ganglion of the rabbit was studied with the use of intracellular electrodes. Caffeine (3–9 mm) enhanced the synaptic transmission through the superior cervical ganglion and evoked in the neurons of the superior cervical ganglion rhythmic hyperpolarizations and sustained hyperpolarization. Aminophylline produced similar effects. Rhythmic caffeine-induced hyperpolarizations appeared with a peak frequency of 7.9 ± 1.2/min, their maximal amplitudes were equal to 15 mV and the durations of each was 3–4 s. The rhythmic hyperpolarization was followed by a decreased membrane resistance and reversed at the potassium equilibrium potential. The average maximal amplitude of the sustained hyperpolarization was 5.6 ± 0.6 mV. In most neurons the latter was associated with an increased membrane resistance, did not decrease or reverse at the potassium equilibrium potential and could be blocked by strophanthin (0.08 mm) or by partial replacement of external sodium with lithium. In the rest of the neurons the sustained hyperpolarization was preceded by a brief depolarization, was accompanied by a drop in membrane resistance, and reversed as the membrane was hyperpolarized. Hexamethonium (0.1 mm) and atropine (0.05 mm) did not block either type of caffeine-induced hyperpolarization.The effect of caffeine (9 mm) on sodium efflux from the ganglion loaded with²²Na⁺ was studied. Caffeine increased sodium efflux from the ganglion and strophanthin (0.08 mm) blocked this effect.It is suggested that caffeine has two effects in the neurons of the superior cervical ganglion (i) rhythmic increases or a sustained increase in membrane potassium conductance; (ii) an increased extrusion of sodium by the electrogenic sodium pump. These effects of caffeine are not mediated by a cholinergic synaptic mechanism.     

Role of the electrogenic Na/K pump in disinhibition-induced bursting in cultured spinal networks

Disinhibition-induced bursting activity in cultures of fetal rat spinal cord is mainly controlled by intrinsic spiking with subsequent recurrent excitation of the network through glutamate synaptic transmission, and by autoregulation of neuronal excitability. Here we investigated the contribution of the electrogenic Na/K pump to the autoregulation of excitability using extracellular recordings by multielectrode arrays (MEAs) and intracellular whole cell recordings from spinal interneurons. The blockade of the electrogenic Na/K pump by strophanthidin led to an immediate and transient increase in the burst rate together with an increase in the asynchronous background activity. Later, the burst rate decreased to initial values and the bursts became shorter and smaller. In single neurons, we observed an immediate depolarization of the membrane during the interburst intervals concomitant with the rise in burst rate. This depolarization was more pronounced during disinhibition than during control, suggesting that the pump was more active. Later a decrease in burst rate was observed and, in some neurons, a complete cessation of firing. Most of the effects of strophanthidin could be reproduced by high K+-induced depolarization. During prolonged current injections, spinal interneurons exhibited spike frequency adaptation, which remained unaffected by strophanthidin. These results suggest that the electrogenic Na/K pump is responsible for the hyperpolarization and thus for the changes in excitability during the interburst intervals, although not for the spike frequency adaptation during the bursts.     

Hyperpolarizing current of the Na/K ATPase contributes to the membrane polarization of the Purkinje cell in rat cerebellum

 The contribution of the Na/K ATPase (pump) current to the polarization of the Purkinje cell has been studied using slices of the rat cerebellum by blocking the pump with dihydro-ouabain (DHO) while recording the membrane potential with microelectrodes in the somata. From our recordings, it appeared that blocking the pump depolarized the Purkinje cells more rapidly than might be expected from shifts in Na⁺ and K⁺ concentrations, suggesting the removal of a hyperpolarizing current. Application of DHO, in the presence of tetrodotoxin (TTX), led to calcium spike firing and plateau-like discharges suggesting activation of voltage-dependent calcium channels in the dendrites. Adding 2 mM Co²⁺ to the medium did not prevent the depolarizations. Removing calcium from the bathing medium containing 2 mM Co²⁺ blocked the spiking activity but DHO application still produced a depolarization. Experiments to measure the current inhibited by DHO indicated that the Na/K pump supplies a constant current of 240 pA. Substitution of the sodium with choline produced a hyperpolarization, during which DHO had no effect on the membrane potential. Substitution of the sodium with lithium produced only a slowly developing depolarization. It is concluded that in the cerebellar Purkinje cell, a continuous sodium ion influx activates the pumps which produce a current that directly contributes to the membrane polarization. Possible pathways for this sodium influx are discussed. Received: 3 April 1997 / Received after revision and accepted: 12 May 1997     

Calcium-dependent potentials in mammalian red nucleus neurons in vitro

Intracellular recordings were made from red nucleus (RN) neurons in guinea-pig slice preparations. The slow afterhyperpolarization (AHP) following an action potential was reversibly abolished by Co2+ or Mn2+. Its amplitude was dependent on the extracellular K+ concentration. When tetraethylammonium was added to the perfusing solution, a tetrodotoxin-resistant regenerative depolarization was evoked which was blocked by Co2+ or Mn2+. There results suggest that the slow AHP is produced by an increase in Ca2+-dependent K+ conductance and that RN neurons have a voltage-dependent Ca2+ conductance.