Fall In Membrane Resistance Research Articles

Actions of vasodilator nerves on arteriolar smooth muscle and neurotransmitter release from sympathetic nerves in the guinea-pig small intestine.

1. Brief constrictions of arterioles of the isolated submucosa of the guinea-pig small intestine were evoked by stimulation of the perivascular sympathetic nerves. Prior stimulation of vasodilator neurones in the submucosal nerve plexus greatly reduced the constrictor response to sympathetic stimulation. 2. Vasodilator nerve stimulation reduced both the amplitude and rate of decay of the excitatory junction potential (EJP) evoked in the arteriolar smooth muscle by sympathetic nerve stimulation. 3. Computer simulation of the effect of membrane resistance changes on the EJP amplitude indicated that the change in amplitude could not be explained by the fall in membrane resistance alone, suggesting that vasodilator nerve activity reduced neurotransmitter release from the sympathetic nerves.

Correlation of anoxic neuronal responses and calbindin-D28k localization in stratum pyramidale of rat hippocampus.

Immunohistochemical staining for the calcium-binding protein calbindin-D28k (CaBP) was combined with Lucifer Yellow (LY) identification and intracellular recording of changes in membrane parameters of pyramidal neurons in CA2, CA1, and the subiculum of rat hippocampal slices during brief exposure (4.0 +/- 0.19 min) to N2. Anoxia evoked either a depolarization or hyperpolarization of membrane potential (VM) (+21.5 +/- 2.79 mV above VM = -70.5 +/- 1.50 mV, n = 30 and -7.2 +/- 0.72 mV below VM = -68.2 +/- 1.34 mV, n = 24, respectively) and a fall in membrane resistance of approximately 20%. Differences in the response could be correlated with the presence or absence of CaBP and the localization of neurons in different layers of stratum pyramidale and sectors of the hippocampus. For neurons immunopositive for calbindin (CaBP(+)), depolarization was observed more frequently (83%) than hyperpolarization (17%); in contrast, 44% of responses of calbindin-negative (CaBP(-)) neurons were depolarizing and 56% were hyperpolarizing. Depolarizations of CaBP(+) neurons were more gradual in slope, and more rapidly reached a plateau in comparison with those recorded in CaBP(-) neurons. Responses of neurons in the superficial layer of stratum pyramidale (in which 79% of CaBP(+) pyramidal neurons were situated) were mainly depolarizing (91%), while for those in the deep layer (which contained 89% of the CaBP(-) cells) such responses were observed less often (45%). Depolarization was also more common than hyperpolarization for cells located in CA2/CA1c/CA1b (63%) than in the CA1a/subicular region (37%). The depolarizing response of the majority of pyramidal neurons which are CaBP(+), superficial, and closer to CA3 may reflect an efficient buffering of intracellular Ca2+, which maintains a low [Ca2+]i, steep gradient for Ca2+ influx and may facilitate the movement of Ca2+ away from points of entry. The neurons which are CaBP(-), deep, and closer to subiculum and in which N2 evokes hyperpolarization, on the other hand, may have a sustained elevation/accumulation of cytosolic Ca2+ which could activate K+ conductance, inhibit Ca2+ influx, and stabilize the membrane potential. These experiments provide a functional correlate for CaBP and suggest that it may have a significant role in Ca2+ homeostasis and the determination of selective neuronal vulnerability.

Hypoxia-activated Ca2+ currents in pacemaker neurones of rat rostral ventrolateral medulla in vitro.

We examined the effects of brief periods of hypoxia or application of cyanide on the discharge and membrane properties of medullary pacemaker neurones in slices of the rostral ventrolateral reticular nucleus (RVL) of the medulla oblongata of rats. Stable intracellular recordings were obtained from seventy-nine neurones within the RVL which exhibited spontaneous rhythmic discharge in the absence of excitatory postsynaptic potentials (EPSPs). The membrane potential cycles of these neurones could be reset with an evoked spike without eliciting EPSPs or inhibitory postsynaptic potentials and hence met criteria of RVL pacemaker neurones. Hypoxia, produced by reducing O2 from 95 to 20% for 40 s or exposure to cyanide (30-300 microM for 40 s), reversibly increased neuronal discharge 1.6-fold (20% O2) or 2.6-fold (300 microM cyanide), respectively, in association with membrane depolarization and a significant fall in membrane resistance. The membrane responses to hypoxia and cyanide were observed in the presence of tetrodotoxin (TTX) at a concentration (10 microM) which eliminated spontaneous spikes or spikes evoked by intracellular depolarization. When recorded at a holding potential of -70 mV by single-electrode voltage clamp, hypoxia or cyanide (300 microM) elicited inward currents of 0.44 +/- 0.06 and 0.58 +/- 0.08 nA, respectively, which are attenuated by reducing the concentration of extracellular Ca2+ ions, and abolished by 2 mM CoCl2 and 100 microM NiCl2, but not affected by 50 microM CdCl2, replacement of 83% extracellular Na+, or adenosine deaminase (2U ml-1). We conclude that hypoxia and cyanide directly excite RVL pacemaker neurones in vitro by a common mechanism: activation of Ca2+ channel conductance.

Calcitonin gene-related peptide evokes distinct types of excitatory response in guinea pig coeliac ganglion cells

Pressure application of calcitonin gene-related peptide (CGRP) evoked in a population of guinea pig coeliac neurons 3 types of response: a fast, a slow and a biphasic depolarization. The responses were not appreciably affected in low Ca/high Mg or tetrodotoxin-containing Krebs solution. The fast depolarization was associated with a fall in membrane resistance; it was made larger on hyperpolarization and the estimated reversal potential was −24 mV. The fast response was reversibly blocked in a Na-free medium as well as by relatively high concentrations of d-tubocurarine (50–100 μM) but not by hexamethonium. The slow, CGRP-induced depolarization resistant to nicotinic and muscarinic antagonists, was associated with either a small increase or decrease of input resistance. Membrane hyperpolarization increased the slow response in the majority of coeliac neurons, with an estimated reversal potential of −44 mV. The biphasic depolarization displayed electrophysiological and pharmacological characteristics resembling the fast and slow responses. These results raise the possibility that CGRP acting via two distinct types of receptor elicits, respectively, a fast, Na-dependent excitatory response and a slow response, the mechanism of which remains to be established.

Fast and slow depolarizing responses of guinea-pig coeliac ganglion cells to 5-hydroxytryptamine

Responses to 5-hydroxytryptamine (5-HT) applied by pressure ejection to guinea-pig coeliac ganglion cells were recorded intracellularly in vitro. Three types of responses were observed: a fast depolarization (43% of cells), a slow depolarization (25%) and a biphasic response comprising an initial fast followed by a slow depolarization (30%). Fast depolarizations had a mean duration of 6.4 s and were associated with a decrease in membrane input resistance. On hyperpolarizing the membrane, responses increased in amplitude; the mean reversal potential was −23 mV. Fast responses were nearly eliminated in a Na-free medium but were unaffected by low Ca/high Mg solution or by low Cl medium. Fast responses were depressed by MDL 72222, quipazine, metoclopramide, and by relatively high concentrations of d-tubocurarine, but unaffected by methysergide. Slow responses had a mean duration of 62 s and were associated either with no change or a slight fall in membrane resistance. On hyperpolarizing the membrane, slow responses were increased in most but not all cells tested. Slow depolarizations were depressed by methysergide but unaffected by agents that suppressed the fast responses. The biphasic response displayed electrophysiological and pharmacological characteristics of both the fast and slow depolarizations. It is concluded that guinea-pig coeliac ganglion cells display either a rapid depolarization, a slow depolarization or a biphasic response to 5-HT, that the fast depolarization appears to be mediated by a conductance increase to Na and K, while a different mechanism appears to underlie the slow depolarization, and that the two responses are mediated by different kinds of 5-HT receptor.

The effect of noradrenaline on the ion permeability of isolated mammalian hepatocytes, studied by intracellular recording.

1. The influence of noradrenaline on the membrane potential and conductance of isolated guinea-pig and rabbit hepatocytes in short-term (2-8 h) tissue culture has been studied by intracellular recording. 2. Resting hepatocytes had linear current-voltage relationships, with input resistances of 166 and 216 M omega in guinea-pig and rabbit cells respectively. The recorded membrane potential was -18 mV in each species, though the true resting potential is likely to have been up to 10 mV greater. 3. The hepatocytes sometimes slowly hyperpolarized during intracellular recording, and this was associated with a fall in membrane resistance, and an increase followed by a decrease in membrane potential noise. These effects were abolished by quinine (200 microM) but not by apamin (50 nM), and are attributable to a K+ conductance activated by cell swelling. 4. Noradrenaline (2 microM, in the presence of propranolol at 1 microM) was applied to individual hepatocytes by pressure ejection (puffer pipette technique). After a short latency, the cells hyperpolarized by a mean of 18 mV in both guinea-pig and rabbit preparations. This was associated with a large rise in membrane conductance (50 nS in guinea-pig, 54 nS in rabbit cells). The reversal potential for this action was -38 mV. 5. The experiments were repeated in the presence of apamin (50 nM) to block the Ca2+-dependent K+ permeability which noradrenaline activates in these cells. Noradrenaline still caused some hyperpolarization and a substantial increase (approximately 40 nS) in conductance, with a reversal potential (Er) of -31 mV. This can be attributed to an increase in Cl- conductance. 6. In keeping with this interpretation, noradrenaline applied in the absence of Cl- (replaced by isethionate or gluconate) caused a much greater hyperpolarization (58 mV in guinea-pig, 40 mV in rabbit cells) associated with a smaller rise in conductance (approximately 12 nS). Er for this action was -95 mV (guinea-pig) and -68 mV rabbit), suggesting that the conductance increase was now mainly to K+. 7. The magnitudes of the conductance changes produced by noradrenaline under the various experimental conditions suggest that the increase in the conductance to Cl- (delta GCl) is 3-fold greater than that to K+ (delta GK). 8. The activation of delta GCl occurs either at the same time as delta GK, or (in ca. one cell in ten) a few seconds later.(ABSTRACT TRUNCATED AT 400 WORDS)

Extracellular calcium and microwave enhancement of membrane conductance in snail neurons.

Microwave irradiation has been shown to decrease the input resistance of snail neurons. In this study, we examined the role of extracellular calcium in triggering the microwave-induced enhancement of membrane conductance. Two sets of experiments were conducted. In the first set, nerve cells were superfused using Ringer solution with added Cd2+ (0.9 mM) which is a known blocker of calcium channels. In the second set, cells were superfused with low Ca2+ (0.7 mM) Ringer solution. Microwave irradiation was conducted at 2,450 MHz for 30 min with a specific absorption rate of 13 mW/g. It was found that 7 mM to 0.7 mM lowering of Ca2+ in bathing solution as well as blocking of calcium channels in neuronal membrane by means of Cd2+ did not influence the fall in membrane resistance induced by microwave radiation. In fact, the observed changed in membrane resistance in these experiments were nearly equal to those observed for neurons superfused by normal Ringer's. Thus, these results rule out the possible contribution of external Ca2+ in the observed microwave effect. Experiments with high Ca2+ solution also support this conclusion.

Evidence of slow IPSP in mammalian prevertebral ganglia

A short train of nerve stimulation evoked in a portion of neurons of the guinea pig inferior mesenteric ganglia a slow hyperpolarization (slow IPSP) which persisted when the preceding orthodromic spikes were reduced to subthreshold EPSPs by curare, was reversibly abolished in a low Ca solution, and was not blocked by atropine; furthermore, the post-tetanic hyperpolarization induced by direct intracellular stimulation was generally smaller than the slow IPSP evoked by nerve stimulation. The slow IPSP was often associated with a fall in membrane resistance and its amplitude decreased with membrane hyperpolarization. The results indicate that the slow hyperpolarization is a synaptic potential, and differs from the slow IPSP observed in mammalian and amphibian paravertebral ganglionic neurons in that it is atropine insensitive.

Post-tetanic depolarization in sympathetic neurones of the guinea-pig.

1. Repetitive intracellular stimulation at a frequency of 5-30 Hz for 1-10 s evoked in neurones of the isolated inferior mesenteric and superior cervical ganglia of the guinea-pig three types of post-spike membrane potential changes: (i) hyperpolarization, (ii) hyperpolarization followed by a slow depolarization, and (iii) a second hyperpolarization following the initial two responses.2. The initial post-spike hyperpolarization had a mean duration of 2.0 s and was often associated with a fall in membrane resistance; it could be elicited in every sympathetic neurone studied. This response was termed the post-tetanic hyperpolarization (PTH).3. The slow depolarization which could be induced only in a portion of neurones had a mean amplitude and duration of 2.2 mV and 27.5 s, respectively; it was termed the post-tetanic depolarization (PTD).4. PTD was associated with a fall in membrane resistance, augmented by membrane hyperpolarization, and reduced by depolarization; its mean extrapolated equilibrium potential was -38 mV.5. PTD was not blocked by nicotinic and muscarinic antagonists, or alpha-and beta-adrenergic receptor antagonists, whereas it was suppressed by adrenaline, noradrenaline, Co(2+) and a low Ca(2+) solution.6. The amplitude of the single spike after-hyperpolarization in normal Krebs solution as well as in high K(+) solution was increased during PTD; furthermore, conditioning hyperpolarization to the level of E(K) increased the amplitude of PTD in normal Krebs as well as in high K(+) solution.7. PTD with similar amplitude, time course and membrane characteristics could be evoked in a portion of neurones of the rabbit superior cervical ganglia; however, PTD was not detected in neurones of the rat superior cervical ganglia.8. Decentralization of the guinea-pig and rabbit superior cervical ganglia for 14 d did not alter the number of neurones in which PTD could be elicited, its amplitude, or its time course.9. Our results suggest that a chemical substance(s) is responsible for the generation of PTD; it may be released from the soma and/or dendrites and acts in an auto-receptive manner on the cells in question. The nature and origin of the second hyperpolarization remain to be clarified.

Membrane properties of Aplysia neurones intracellularly injected with phospholipases A and C.

1. The effects of phospholipases A from bee venom and from porcine pancreas and of phospholipases C from Clostridium welchii and Bacillus cereus on active and passive membrane properties of Aplysia neurones have been studied. Consistent alterations in electrical membrane properties were found following intracellular application of three of these enzymes.2. Bee venom phospholipase A produced a rapid decrease of membrane potential and resistance. Voltage clamping revealed a marked depression of peak transient current with little or no effect in the late outward current.3. Mammalian phospholipase A was found ineffective in changing either the resting or active membrane properties.4. Phospholipase C from Bacillus cereus led to a strong hyperpolarization and a fall in membrane resistance. Voltage clamping revealed a marked increase in the late outward current.5. Neurones injected with Clostridium welchii phospholipase C manifested a several-fold rise in resting membrane resistance as well as a tendency to slight hyperpolarization.6. All enzymes were ineffective when externally applied.7. It is tentatively concluded that the internally applied phospholipases affect specific ionic permeabilities both in the resting and active excitable membrane. Various mechanisms by which the differing actions of enzymes of the same type could be explained are discussed.

Effects of morphine on myenteric plexus neurones

Intracellular recordings were made from neurones in the isolated myenteric plexus of the guinea-pig ileum. Approximately 50% of neurones responded to bath application of normorphine (10 nM−1 μM) with an immediate hyperpolarization. The hyperpolarization was usually associated with a fall in membrane resistance. The effect reversed on washing out the normorphine and was prevented by the addition of the opiate receptor antagonist naloxone (50–100 nM). It is suggested that the membrane hyperpolarization may be the basis for the inhibitory effects of morphine on neurone firing rates which have been reported both in the myenteric plexus and the central nervous system.

The sensitivity of Helix aspersa neurones to injected calcium ions.

1. When calcium chloride was injected into Helix aspersa neurones there was a fall in membrane resistance and the membrane potential became hyperpolarized.2. The reversal potential of the response was dependent on the concentration of potassium in the external solution.3. Injection of a calcium-EGTA buffer containing 9 x 10(-7)M free calcium reduced the membrane resistance by 25%. When calcium chloride was injected it was necessary to increase the total intracellular calcium concentration by about 10(-3)M to produce similar change of resistance.4. In sodium-free (Tris) solution there was a slow fall of membrane resistance as if the intracellular calcium concentration had increased. There was a similar resistance change in the presence of 2,4-dinitrophenol and iodoacetate.5. A series of repetitive depolarizing pulses produced a long lasting reduction in membrane resistance which was enhanced by 2,4-dinitrophenol and iodoacetate.6. It is concluded that (a) injection of calcium causes an increase in potassium permeability, (b) the injected calcium is rapidly pumped from the cytoplasm by a sodium-dependent mechanism and by mitochondria, and (c) 1-2 msec depolarizing pulses stimulate an influx of calcium. This influx is rapid enough to trigger potassium activation during an action potential.

Effects of electrogenic sodium pumping on the membrane potential of longitudinal smooth muscle from terminal ileum of guinea-pig.

1. The membrane potential of the separated longitudinal muscle of the guinea-pig terminal ileum was recorded intracellularly with glass micro-electrodes.2. In tissues kept at room temperature and then brought to 35 degrees C for 15-30 min or about 1 hr, the fall in membrane potential upon changing to potassium-free solution was 21.4 +/- 3.5 mV and 13.4 +/- 1.8 mV respectively. Ouabain (1.7 x 10(-6)M) produced a fall in membrane potential of 8.1 +/- 1.1 mV. Returning potassium to potassium-free solution, or changing from ouabain-containing to ouabain-free solution, resulted in an increase in membrane potential which was greater than the initial fall.3. Readmitting potassium to potassium-free solution produced an increase in membrane potential which began within 10 sec and reached a maximum within 15-30 sec. This response was reduced, abolished, or converted to a depolarization by ouabain. In chloride-deficient (13 mM) solution in which membrane resistance was increased, the response to readmitting potassium was increased 2(1/2)-fold so that the membrane potential sometimes exceeded -100 mV, which was probably more negative than E(K). On the basis of these results it was assumed that the response to readmitting potassium was due to the electrogenic activity of the sodium pump.4. The response to briefly readmitting a fixed concentration of potassium increased during the first 30 min in potassium-free solution. This increase was not due to an increase in membrane resistance as this fell with time in potassium-free solution. It was suggested that the increase in the response resulted from the progressive rise in internal sodium concentration which is known to occur in smooth muscle in potassium-free solution.5. Increasing the concentration of potassium over the range approximately 0.1-20 mM, increased the size of the electrogenic potential observed upon readmitting potassium to potassium-free solution. There was a fall in membrane resistance upon readmitting potassium (0.6, 5.9, or 20 mM) which was greater the larger the concentration of potassium. When allowance was made for the fall in membrane resistance, the dependency of the electrogenic response upon the concentration of potassium over the range 0.6-20 mM was much increased.6. The results indicate that the rate of electrogenic sodium pumping in this tissue is increased by increasing the external potassium concentration, and probably by increasing the internal sodium concentration. It was suggested that a rise in the latter could sensitize the pump to an increase in the former.

Injections of calcium ions into spinal motoneurones.

1. In cats under Dial anaesthesia, Ca(2+) was injected inside lumbosacral motoneurones, by passing currents between CaCl(2)- and KCl-containing barrels of compound micropipettes.2. There was a reduction in excitability and a fall in membrane resistance, both rapid in onset and quickly reversible.3. The minimum effective injection current was approximately 10 nA, and the effect reached a maximum with currents of approximately 30 nA. The mean slope of resistance change against injection current was -1.7%/nA (S.E. 0.35).4. The most common change in membrane potential was a hyperpolarization; but in nearly half the cases, there was no clear change or a small depolarization. A reversal level for the effect of Ca(2+) could be measured in five cells: on the average, it was 10 mV more negative than the resting potential.5. Observations on i.p.s.p.s showed that Ca(2+) probably does not alter g(Cl): it was concluded that the fall in membrane resistance caused by intracellular Ca(2+) is mainly due to an increase in g(K).6. These results confirm previous suggestions that a steep transmembrane gradient of Ca(2+) is essential for the maintenance of a low membrane conductivity, and that a rise in internal free Ca(2+) - whether due to influx or release from internal stores - may play an important role in regulating neuronal activity.

Actions of dinitrophenol and some other metabolic inhibitors on cortical neurones.

1. In cats under methoxyflurane, DNP and other metabolic inhibitors were tested on cortical neurones by iontophoresis from micropipettes.2. DNP, dinitro-o-cresol, iodoacetate, pentachlorophenol and oligomycin (uncouplers or inhibitors of oxidative phosphorylation), as well as moderate anoxia, blocked selectively and reversibly spontaneous firing and discharges evoked by ACh; responses evoked by glutamate were facilitated by moderate doses of DNP and blocked only by large amounts.3. Azide, cyanide, ouabain and strophanthidine had a mainly excitatory effect; the cardiac glycosides tended to depress more strongly responses to glutamate.4. Intracellular observations showed that DNP causes a sharp fall in electrical excitability, associated with a hyperpolarization and fall in membrane resistance.5. The hyperpolarizing action of DNP had a mean reversal level (E(DNP)) nearly 30 mV more negative than the resting potential; E(DNP) was identical with the mean reversal level for the depolarizing action of ACh, measured on the same cells.6. DNP had its usual hyperpolarizing effect on neurones whose IPSPs had been made positive by raising the internal [Cl]; the mean E(IPSP) was over 30 mV more positive than E(DNP).7. It is concluded that DNP lowers excitability by raising the membrane conductance to K(+) (g(K)) and that it blocks ACh responses selectively because ACh has a precisely opposite action on these neurones.8. In the Discussion, it is suggested that the rise in g(K) is mediated by an increase in internal free Ca(2+), caused by a slowing of mitochondrial activity, and that a similar mechanism may play a significant role in general anaesthesia.

Some properties of unresponsive cells in the cerebral cortex.

Unresponsive cells in the pericruciate cortex of cats under pentobarbitone were distinguished from neurones by total absence of spontaneous or evoked spikes and synaptic responses. They had much greater membrane potentials (mean −62 mV) and resistances (mean 48 MΩ) than neurones (means −36 mV, 21 MΩ), and the potentials were unusually stable. They could be depolarized by external applications of K+, but were insensitive to internal injections of K+ or Cl−. Unlike neurones, they were not hyperpolarized by GABA or depolarized by L-glutamate, but large doses of GABA had a depolarizing effect, without reducing the membrane resistance. ACh also depolarized some neurones and unresponsive cells without a fall in membrane resistance; but nor-adrenaline and 5HT were ineffective on the few unresponsive cells tested. It is suggested that unresponsive cells are neuroglia; that the depolarizing actions of GABA and ACh indicate a change in electrogenic active transport, and that glia may remove transmitters from the extracellular space.

The effects of hypoxia on neuromuscular transmission in a mammalian preparation.

1. The rat diaphragm-phrenic nerve preparation in vitro failed to contract in response to nerve impulses after 10-20 min exposure to solutions containing 95% N(2) and 5% CO(2) (hypoxic solutions) at temperatures between 33 and 38 degrees C. Intracellular recording revealed that end-plate potential (e.p.p.) amplitudes fell below the firing threshold for muscle fibres and then disappeared probably because of block of intramuscular nerve conduction.2. In curarized and Mg-paralysed preparations the reduction in e.p.p. amplitudes was found to be due to a fall in their quantal content. In about half of the Mg-paralysed preparations, however, and in curarized preparations after repeated exposures, there were increases in quantal content of e.p.p.s during hypoxia.3. Miniature end-plate potential (m.e.p.p. frequency increased in a cyclic fashion during hypoxia and this increase was largely suppressed in the presence of a raised extracellular Mg concentration. M.e.p.p. amplitude increased (range 0-100% of control value) after about 20 min hypoxia.4. Post-tetanic potentiation of e.p.p. amplitudes and m.e.p.p. frequency was reduced after exposure to hypoxic solutions. During hypoxia the e.p.p. amplitude potentiation was reduced but the m.e.p.p. frequency potentiation was augmented.5. There was an increase in the post-synaptic sensitivity to carbamylcholine after 20 min hypoxia which was sufficient to explain the increase in m.e.p.p. amplitude. Other post-synaptic changes were a fall in membrane potential (average 6 mV after 20 min) and a fall in membrane resistance after 30-60 min exposure to hypoxia.6. The effects of hypoxia upon neuromuscular transmission were partially explained by reduction of active transport of sodium and potassium ions and consequent depolarization of nerve and muscle.