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Although KATP channels have been proposed as playing a role in most types of myocardial damage associated with ischemia/reperfusion, the potential benefits of KATP channel modulators against the biochemical and electrical disturbances observed during ischemia remain unclear. We have thus studied the effects of glibenclamide and cromakalim, KATP channel blocker and opener respectively, on membrane lipid injury and arrhythmias, in a model of ischemic-reperfused guinea-pig myocardium. Ventricular strips were prelabeled with [3H] arachidonic acid, then subjected to normal conditions (Time-related Control) or to simulated ischemic-reperfused conditions in absence of drug (Control) or in presence of glibenclamide 10 microM or cromakalim 10 microM. The release of radioactive compounds was counted by liquid scintillation spectrometry, while action potentials (AP) were recorded with intracellular microelectrodes. Reperfusion induced a significant increase of arachidonic acid release (P0.05 versus Time-related Control). Glibenclamide inhibited the reperfusion-induced arachidonic acid release while cromakalim only delayed it (respectively 483 +/- 87 dpm/g, P0.05 and 790 +/- 143 dpm/g. NS versus 838 +/- 80 dpm/g for Control, after 30 min of reperfusion). Unlike glibenclamide, cromakalim was proarrhythmic during reperfusion (in 100% of preparations versus 33% in Control or in presence of glibenclamide, P0.05). This in vitro study shows that glibenclamide prevented the reperfusion-induced membrane arachidonic acid release, without proarrhythmic effect, whereas cromakalim, associated with proarrhythmicity, was unable to protect myocardium from cell lipid damage.

We used intracellular microelectrodes to study the effects of histamine on both normal and abnormal automaticity in sheep Purkinje fibers. Histamine, dimaprit and 4-methylhistamine caused a similar reduction of the action potential duration in driven Purkinje fibers. Histamine (10-6 M) induced spontaneous activity in p previously quiescent preparations more often than did equimolar concentrations of dimaprit and 4-methylhistamine. The effects of histamine on automaticity were enhanced in the presence of barium In fact histamine, at concentrations which were unable to induce automaticity in normal preparations, induced it in the presence of barium. In Purkinje fibers manifesting barium-induced automatic activity, histamine (10-7--10-6M) significantly increased the average number of spontaneous action potentials and shortened the time of their appearance. In the same range of concentrations, histamine dose-dependently increased the iological manifestation of calcium overload. Histamine (10-6--10-4M) increased the OAP amplitude of strophanthidin -treated Purkinje fibers, eventually inducing triggered extrabeats. All these previously described effects were selectively blocked by cimetidine (10-5 M). It is concluded that histamine may induce cardiac arrhythmias under conditions of calcium overload and that this effect may be due to induction or enhancement of oscillatory afterpotentials.

We studied the actions and interactions of ethanol and imipramine on the sinus node. Strips of the right rat atrium including the sinus node were superfused with Tyrode's solution at 37°C while beating spontaneously. The preparations were exposed to imipramine or ethanol alone as well as to the two drugs in combination while recording membrane potentials with standard intracellular microelectrodes. The results obtained show that ethanol 0.8 and 2.4 g/l exerted a positive chronotropic action. On the other hand, imipramine 0.25 mg/l did not modify the sinus node rate. However, it reduced significantly the positive chronotropic action of ethanol. The sinus node rate decreased under the action of a higher concentration of imipramine (1 mg/l). When ethanol was tested in combination with this concentration of imipramine, the effect of the latter prevailed. In conclusion, a concentration of imipramine that did not affect the sinus node rate antagonized the positive chronotropic action of ethanol. In addition, the negative chronotropic action of a higher concentration of imipramine prevailed over the positive action of ethanol. The results obtained provide additional support to the notion that the use of ethanol and cardioactive drugs in combination may result in significant changes in the actions of either of the two, or both. This is of clinical relevance, since at least some of the individuals under treatment with cardioactive drugs will be alcoholics and/or social drinkers.

The physicochemical characteristics of the terbium-adriamycin complex (terbomycin) were studied. Perturbations in the visible absorption spectrum of adriamycin by terbium (Tb3+) was indicative of formation of the terbomycin complex. The absorption maximum of free adriamycin at 479 nm shifted towards the absorption maximum of terbomycin at 539 nm. The binding of Tb3+ to adriamycin was negligible at acidic pH. At alkaline pH, the affinity of Tb3+ for adriamycin increased. The stoichiometry of binding was estimated to be 0.5; one Tb3+ ion per two adriamycin molecules. Thermodynamic analysis revealed that the spontaneous formation of terbomycin was due to an increase in the entropy of the system. The effects of adriamycin, Tb3+ and terbomycin on sinus node automaticity were studied using sinus node from rats, superfused with modified mammalian Tris-Tyrode's solution (37 degrees C). The sinus node rate was monitored with intracellular microelectrodes. 25 microM Tb3+ increased the sinus node rate. Adriamycin (50 microM) depressed sinus node automaticity. Terbomycin also reduced the sinus node rate. There was no difference between the effects of adriamycin and terbomycin. The chronotropic effect of terbomycin persisted in the presence of atropine.

The effects of lindane, a gamma-isomer of hexachlorocyclohexane, were studied on transmembrane potentials and currents of frog atrial heart muscle using intracellular microelectrodes and the whole cell voltage-clamp technique. Lindane (0.34 microM to 6.8 microM) dose-dependently shortened the action potential duration (APD). Under voltage-clamp conditions, lindane (1.7 microM) increased the amplitude of the outward current (Iout) which developed in Ringer solution containing TTX (0.6 microM), Cd2+ (1 mM) and TEA (10 mM). The lindane-increased Iout was not sensitive to Sr2+ (5 mM). It was blocked by subsequent addition of quinidine (0.5 mM) or E-4031 (1 microM). E-4031 lengthened the APD; it prevented or blocked the lindane-induced APD shortening. In conclusion, our data revealed that lindane increased the quinidine and E-4031-sensitive rapid delayed outward K+ current which contributed to the AP repolarization in frog atrial muscle.

1. Bovine ventricular myocytes isolated according to isenberg and Klockner (1982a) were superfused with Tyrode solution ([Ca]0-3.6 mM, 35°C) and investigated with intracellular microelectrodes. The properties of the action potential (AP) were compared to those published for multicellular ventricular tissue. 2. At 1 Hz stimulation, the AP had 4 clearly discernible phases with a trapezoidal configuration and a duration of 276 ±38 ms. At [Ca]0=1.8 mM the 4 phases were less clear. Following a 1 min rest period, 1 Hz stimulation shortened the AP by 28% ([Ca]0=3.6 mM) or by 12% ([Ca]0=1.8 mM). 3. The AP started with a sodium spike which rose with\(\dot V_{\max }\) of 250±60 V/s to an overshoot of +40±5 mV. The sodium spike was blocked by 60 μM TTX.\(\dot V_{\max }\) was modified with 2s conditioning pre-pulses; the resultingh∞ curve showed half maximal inactivation atV0.5=−65 mV and a slope factork=5.5 mV. Reduction of [Ca]0 to 1.8 mM shiftedh∞ by −5 mV but did not modifyk. However, large shifts inh∞ curves were measured in myocytes depolarized by 20 mM or 0 mM [K]0. 4. Slow AP's were studied when the sodium spike was inactivated by high [K]0 or blocked by TTX. The slow AP's rose with\(\dot V_{\max }\) of 11±5 V/s to a maximum plateau potential of 36±4 mV;\(\dot V_{\max }\) increased up to 24 V/s when the latency was shortened by increased stimulus strength.\(\dot V_{\max }\), plateau and duration were increased by 0.1 μM adrenaline, reduced by 1.8 mM [Ca]0 and blocked by Ni, Co or D600. 5. Changes in [K]0 modified the resting potential (Vm) as discribed in multicellular ventricular tissue.Vmax was −84 ±4 mV at [K]0=5.4 mM and −54±3 mV at [K]0=20 mM; the latter value suggests [K]i=152±8 mM. The increase in [K]0 abolished the sodium spike and shortened the AP, but did not change the maximum plateau potential. In voltage clamp experiments the i-v relation of the repolarizing late outward currents was N-shaped. The sensitivity of these currents to [Cs]0 and [K]0 indicated that the inward rectifying potassium conductance declines with [K]0. 6. The experimental interventions used in the present study changed the AP of the isolated myocyte in a way similar to that reported for multicellular ventricular tissue, i.e. no electrophysiological peculiarity was observed. We conclude that the enzymatic isolation procedure did not significantly alter the properties of the Na-, Ca- or K-channels.

1. The relative spectral sensitivities of identified large second order neurons (L neurons) of the dragonfly ocellus have been examined with dye filled intracellular microelectrodes placed in the brain. 2. Representative L neurons of the four geometric classes present in the median ocellar nerve show essentially similar waveforms in response to white, green and UV light. 3. Details of the retinal projections of these four neuron classes have been established. 4. L neurons exhibit a marked reverse Purkinje shift at low light intensities. Green sensitivity (485 nm) decreased relative to UV sensitivity (360 nm) as the intensity of illumination increased. At a mean intensity of 1.78 × 1012 photons cm−2 s−1 the L neurons are equally sensitive to UV and green light. 5. The reverse Purkinje shift is discussed in relation to data previously obtained for dragonfly receptor cell spectral sensitivity and in relation to its possible functional significance.

The construction and use of tissue holders that allow simultaneous long-term recordings of mechanical and intracellular electrical activity from a variety of smooth-muscle preparations is described. The tissue holders permit adjustment of tissue stretch following its fixation for recording purposes. By this means intracellular microelectrodes can be kept in position despite marked changes in the mechanical activity of the tissue. Since simultaneous recordings of electrical and mechanical activity of up to 2 hrs duration can be made, studies of the relationships between the membrane potential changes and mechanical changes evoked by drugs and neurotransmitters are greatly facilitated.

Isolated sartorius nerve-muscle preparations from Rana temporaria were irrigated in a chamber. Resting muscle membrane potentials and evoked end-plate potentials were recorded via intracellular microelectrodes. Multiple end-plate potentials appeared after the preparation had been exposed to a suspension, in Ringer's solution, of the toxic synthetic pyrethroid: 5-benzyl-3-furylmethyl (+)-cis-chrysanthemate (NRDC 119). It is thought that this effect was due to repetitive activity arising in the motor nerve fibers. The pyrethroid had no effect on the muscle resting potential nor on the amplitude or time course of the initial evoked end-plate potential.

1. E.p.s.p.s and terminal potentials (t.p.s) produced by the action of single medial gastrocnemius Ia afferent fibres were examined with intracellular microelectrodes in cat triceps surae motoneurones. 2. Simple terminal potentials (t.p.s) appeared as positive-negative diphasic waves with a single positive peak. Shape indices of excitatory post-synaptic potentials (e.p.s.p.s) recorded in conjunction with simple t.p.s indicate that these e.p.s.p.s were generated at an electrotonically confined compartment on the motoneurone somadendritic membrane. 3. Compound t.p.s. were similar ones, except that they had two or more positive peaks. Shape indices of e.p.s.p.s recorded in conjunction with compound t.p.s indicate that these e.p.s.p.s were generated at two or more electrotonically separate compartments. 4. E.p.s.p. latency and post-synaptic electrotonic delay were determined for a group of single fibre e.p.s.p.s with simple t.p.s. The shortest recorded latency was 0.26 msec. By subtracting estimated electrotonic delay, the shortest synaptic delay obtained for a medial gastrocnemius Ia single fibre e.p.s.p. was 0.17 msec. 5. The finding of a significant synaptic delay supports the hypothesis that the Ia-motoneurone synapse is a chemically mediated synapse.

(1) Depolarizing after-potentials (DAPs) of hippocampal pyramidal cells were studied with intracellular microelectrodes in rabbits anesthetized with Nembutal and, in addition, curarized. (2) DAPs were classified into two types. One was linearly related in magnitude to the spike and the membrane potential of the cell body while the other was not. Therefore, the former was tentatively called the true DAP and the latter the pseudo-DAP. (3) In accordance with reduction in the membrane potential, the magnitude of the true DAP diminished and at a membrane potential of −60 to −50 mV, the true DAP disappeared. On further reduction in the membrane potential, a hyperpolarization-going dip appeared in place of the true DAP. (4) The pseudo-DAP usually appeared when the membrane potential of the cell body was reduced beyond −60 to −50 mV. But the pseudo-DAP was not quantitatively related to the membrane potential. It was shown that the pseudo-DAP was a nonsynaptic process, and its possible relation to the dendritic spike was discussed. (5) It was demonstrated that it was the pseudo-DAP that led to the genesis of the so-called inactivation response,i.e. a large and prolonged depolarization of the cell membrane with a strong tendency to spike inactivation.

The action of veratridine on the resting membrane potential of intact crayfish axons in the presence of different extracellular alkaline-earth cations has been studied using intracellular microelectrodes. At extracellular pH values of 7, 6 and 5, the apparent binding sequence was consistently Ca2+ ⩾Sr2+ >Mg2+ ≈Ba2+. This particular selectivity sequence and the insensitivity of binding order to changes in extracellular pH have been noted for other biological preparations and provide structural information about the veratridine - interaction sites to which the alkaline earths bind.

Electrocardiograms, ventricular monophasic action potentials registered with suction and intracellular microelectrodes, and pressure curves were recorded simultaneously in isolated rabbit hearts perfused with oxygenated Krebs-Henscleit solution containing varying calcium/magnesium ratios. The duration of the action potential and Q-T interval was not affected by low and high magnesium concentrations unless calcium was omitted or significantly reduced. When Mg was completely absent, solutions free of calcium or with one-sixteenth of the "normal" Ca concentration caused a progressive prolongation of the action potential and Q-T interval until they occupied the entire cardiac cycle. When Mg concentration was low (0.075 to 0.6 mM/L.), the prolongation occurred at a slower rate and was less pronounced. With higher Mg concentrations (1.2 to 19.2 mM/L.), the initial lengthening of the action potential and Q-T interval was followed by a gradual shortening. The higher the Mg concentration employed, the shorter was the period of the initial lengthening, and the more pronounced was the subsequent shortening. These observations can be best explained by assuming that calcium-like action of Mg on the plateau of the action potential becomes apparent only when Ca is absent or significantly decreased. The depression of contractility by calcium deficiency proceeded at the same rate at all studied Mg concentrations. The findings are discussed in relation to the electrocardiographic patterns encountered in clinical cal cium and magnesium deficiency.

The mode of action of tetrodotoxin on the frog muscle fiber membrane has been analyzed with the aid of intracellular microelectrodes. Tetrodotoxin of 10–7 concentration made the applied cathodal current ineffective in producing action potential, whereas the resting potential and resting membrane resistance underwent little or no change. With 10–8 tetrodotoxin the muscle fibers responded with the small action potentials at high critical depolarizations. These results can be explained on the basis of the membrane being stabilized by inactivation of the sodium-carrying mechanism. Although delayed rectification was not observed in normal muscle fibers, it became apparent in the fibers rendered inexcitable by tetrodotoxin. This finding, together with other evidence in the existing literature, supports an applicability of the sodium theory to the frog muscle fibers.

DEVELOPLTENT of electrophysiological technique has provided a means for direct determination of the site of neutropic drug action at cellular level. Intracellular microelectrodes are effective tools for investigation of the mechanism of action of substances affecting synaptic and junctional transmission. The experimental results presented here are concerned with the mechanism of conduction in the neuro-muscular junction.

Effects of Carcholin, Mecholyl, neostigmine and strychnine on spontaneously beating isolated rabbit auricles were studied by recording the transmembrane potential changes with intracellular microelectrodes and the contractions by means of a sensitive mechano-transducer. Electrical recordings were taken in both pacemaker and nonpacemaker areas. Carcholin (10–5 m) and Mecholyl (10–4 m) consistently shortened the duration of the action potential and diminished the developed tension. At higher concentrations Carcholin (10–3 m) and Mecholyl (10–2 m) arrested all auricular activity. Atropine (10–3 m) and epinephrine (10–3 m) restored activity. Following atropine or epinephrine, the inhibitory capacity of the anticholinesterases was abolished. Neostigmine and strychnine had variable effects.

1. The action of three thiamine antagonists (Neopyrithiamine, Oxythiamine, Amprolium) have been studied with intracellular microelectrodes on isolated heart tissue of rat, calf and sheep.

Intracellular microelectrodes have been used to study the site of origin and direction of spread of activity in the rabbit atrium. In this study a number of fiber groups have been found which possess specialized electrophysiological characteristics and a consistent anatomical localization. A pacemaker potentiality has been found only in tissues derived from embryologically distinct structures such as the sinus venosus, the venosus valves and the lower segment of the auricular canal. Activity normally spreads slowly from the S-A node and excites the crista terminalis along a broad front. Spread is then rapid through the crista, the pectinate muscles and the fibers of the atrial roof. The septum is normally excited from the crista terminalis. Excitation reaches the A-V node almost simultaneously from the crista terminalis and the right segment of the S-A ring bundle. This latter shows many characteristics of specialized conducting tissue. Around the A-V ring conduction velocity slows markedly in fibers which have many of the electrophysiological characteristics of A-V nodal fibers.

Potentials of motoneurons of the lower segments of the spinal cord were recorded with the aid of intracellular microelectrodes in experiments on cats with induced tetanus produced by injection of tetanus toxin (1500–2000 mouse LD50) into the extensor muscles of the left shin. Neither afferent volleys of impulses in cutaneous and muscle nerves, nor antidromic volleys in the corresponding ventral roots, produced IPSPs in motoneurons of the extremity into which toxin was injected. The form both of antidromic peak potentials and of monosynaptic EPSPs in motoneurons in which IPSPs were blocked by tetanus toxin did not differ from the form of corresponding potentials of motoneurons in normal cats. The values of threshold depolarization for peak discharges during synaptic and direct stimulation were equal in tetanus and control motoneurons. Resistance and time constant values of the membrane in "tetanus" motoneurons did not differ from the corresponding values for "control" motoneurons.

1. Differences between responses of phasic and intermediate muscle fibers in frogs to prolonged depolarization by a current of threshold strength were investigated with intracellular microelectrodes. 2. Intermediate fibers have a greater tendency toward repetitive spike responses to prolonged depolarization than phasic fibers. This is expressed by the fact that the percentage of fibers responding to threshold depolarization by a series of APs (instead of single APs) is much higher in the group of intermediate, fibers than in the phasic group. In addition, the duration of the repetitive response to depolarization was always greater in intermediate fibers. 3. Agents such as calcium ions and procaine, which depress rhythmic activity in both phasic and intermediate fibers, differ in their quantitative action on these two groups of fibers: concentrations of agents which completely suppress the rhythmic response in phasic fibers merely shorten the duration of the rhythmic discharge in intermediate fibers.

Summary In experiments carried out on isolated neuro-muscular preparation of the frog the action of drugs on junctional transmission was studied with the aid of multi-barrelled intracellular microelectrodes. It was found that different pharmacological agents produce neuromuscular block affecting different parts of the junction : presynaptic nerve terminals (CoCl 2 and Cd Cl 2 ), subsynaptic membrane (curare-like substances), electrically excitable membrane of muscle fibre (narcotics, Ca excess). Some drugs (novocaine) can depress conduction in all parts of the junction.

The effects of the chlorinated hydrocarbon insecticides, DDT and dieldrin, on the electrical activity of skeletal muscle fibres of the clawed toad, Xenopus laevis , were investigated by means of intracellular microelectrodes. Both compounds caused an increase in the amplitude of the negative afterpotential and a slowing-down of the falling phase of the action potential, without affecting the resting potential. Neither DDT nor dieldrin caused repetitive activity. It is concluded that the increase in the duration of the action potential, measured at a potential level of −50 mV, may be responsible for part of the effects of these insecticides on the mechanical response of skeletal muscle.

The electrical “cable” properties of fibers in the soleus muscle of normal and dystrophic mice were measured in vitro at 37C with intracellular microelectrodes. The motor nerve of the dystrophic muscle was stimulated to determine whether or not the fibers chosen for measurement were functionally innervated. Normal and dystrophic muscles which had been surgically denervated were also studied. Specific membrane resistance was higher in normal than in dystrophic muscle fibers; the respective mean values were 1516 and 636 ohm cm 2 . Dystrophic fibers which were not functionally innervated had significantly higher resistance than those which were innervated, but in neither case did values approach the normal. In both normal and dystrophic muscles, previous sectioning of the motor nerve led to an increase in muscle fiber input resistance. Surgically denervated dystrophic fibers had considerably higher resistance than measured in fibers which had become functionally denervated during the course of the disease. The results suggest that denervation per se cannot account for the differences between normal and dystrophic muscle fibers.

Kolmodin, G. M. and C. R. Skoglund. Influence of asphyxia on membrane potential level and action potentials of spinal moto- and interneurons. Acta physiol. scand. 1959. 45. 1–18. — Asphyxial changes in spinal moto- and interneurons of decapitate cats were studied by means of intracellular microelectrodes. In both types of neurons the first change in membrane potential was usually observed after about 30 sec in the form of a depolarization of 5–10 mV which after some 30 sec was followed by a phase of more rapidly developing depolarization of larger amplitude; during recovery a phase of increased polarization may be observed. Interneurons showed signs of being more resistant to asphyxia than moto-neurons. The spib potential underwent a. successive diminution in amplitude, at first owing to the decrease in membrane potential value, later also due to a change in spike peak level. During the later stage of asphyxia a decrease of both the rate of rise of the ascending and the rate of decay of the descending spike phases could be demonstrated. Rhythmical interneuron discharges showed periods of increased and decreased frequency before final blocking.

The effects of varying the potassium and sodium concentrations in the bathing fluid on the resting and action potentials of the cockroach giant axon have been examined by means of intracellular microelectrodes. Plotting the resting potentials against the logarithms of the external potassium concentrations resulted in a straight line in the concentrations higher than about 20 mM. The decrease in the sodium concentration had no appreciable effect on the resting potential, but effectively depressed the action potential. Plotting the reversed potentials against the logarithms of the external sodium concentrations resulted in a straight line, which diverged only slightly from the theoretically expected straight line having a slope of 59 mV for a tenfold change in sodium. The rate of rise of the action potential was decreased very effectively by the sodium-deficient solution. It can safely be said that the overall result constitutes strong evidence for the essential validity of the sodium theory in the cockroach giant axon.

Caudate neurons were recorded with intracellular microelectrodes in halothane anaesthetized cats. Excitatory amino acids and some of their antagonists were applied by microiontophoresis. The agonists exhibited two types of responses, both of which consisted of gradual membrane depolarizations; however, in type 1 this was associated with repetitive, relatively regular firing, whereas in type 2 action potentials generally occurred on additional abrupt depolarizations reaching amplitudes of more than 20 mV and which lasted up to 500 ms.

The effects of temperature on the inhibitory junctional potential (i.j.p.) and the electrotonic potential, recorded in the circular smooth muscle of the guinea-pig ileum, were studied with intracellular microelectrodes. The amplitude and time course of the i.j.p. were both dependent on the ambient temperature, the i.j.p. becoming smaller and more prolonged as the temperature was lowered. In contrast, the electrotonic potential was not much affected by temperature. The atropine-resistant 'late' depolarization was also dependent on temperature. These results are consistent with the mechanism of release, of the nonadrenergic noncholinergic inhibitory transmitter, being sensitive to temperature and with the inhibitory conductance change, underlying the i.j.p., being long compared to the membrane time constant of the circular smooth muscle.

This article briefly reviews the electrical attributes of nerve terminals and of their relationship to secretion for a neurosecretory system in crabs termed the X-organ sinus gland (XOSG). This neurosecretory system is a purely peptidergic secretory system whose hormonal secretions are known to influence or govern nearly all physiologic and homeostatic processes (reviews, Cooke and Sullivan, 1982; Keller, 1983). A portion of the system termed the sinus gland (SG) is a discrete spherical (c. 1.5 mm. diameter) structure composed of an extensive arborization of axon varicosities and terminals together with glial and supportive cells which surround a network of hemolymph sinuses. The cell soma, where axons project to and compose the sinus gland, are located in a cluster named the X-organ. The relatively large size of individual axon terminal dilatations in certain species of crabs (e.g., Cardisoma carnifex. Podoohthalamus vigil) is novel among peptidergic neurosecretory systems. The terminals in these species are of sufficient size (up to 30 um) to have allowed extensive investigation with intracellular microelectrodes in both in situ and in vitro preparations (reviews, Cooke and Stuenkel, 1985; Stuenkel and Cooke, 1988). Recently a preparation of dissociated terminals from the sinus gland has been developed and investigated with the various recording configurations of the patch clamp technique (Lemos et al., 1986). A number of electrical and secretory attributes of the peptidergic terminals have been characterized including: 1) the ionic dependence of regenerative activity, 2) ‘whole terminal’ patch clamp analysis of voltage dependent macroscopic ionic currents and of two novel cation channel types and 3) correlation of terminal electrical behaviour with hormonal secretion monitored simultaneously under different secretory regimes.

The actions and interactions of dihydropyridines (DHPs) and ethanol on sinus node (SN) automaticity were studied using rat sinoatrial preparations superfused with Tyrode's solution at 37 degrees C. Intracellular microelectrodes were used to monitor SN rate (SNR). The automaticity of the SN was not affected by nimodipine 10(-10) M, but it was depressed by a higher concentration of this calcium antagonist (10(-8) M). The racemic compound (+/-)Bay K 8644 had a dose-dependent biphasic chronotropic action: At 10(-8) M it decreased the SNR, while at 10(-7) M it increased the SNR. The negative component was blocked by atropine. A small concentration of ethanol (1.8 x 10(-2) M) had a positive chronotropic effect on the SN, and this action was blocked by nimodipine 10(-10) M. This small concentration of ethanol did not modify the chronotropic effects of either of the DHPs. A higher concentration of ethanol (5.3 x 10(-2) M) did not by itself affect the SNR, but it antagonized the chronotropic actions of the two DHPs.

The ability of dystrophic soleus muscle fibers to generate action potentials in response to supramaximal nerve stimulation was studied with intracellular microelectrodes in mice of the Bar Harbor 129- ReJ dy strain. In experiments conducted in vivo at either 23 or 37 C, about 55% of the fibers examined were abnormal in their ability to generate action potentials. Some of these fibers showed no detectable response to stimulation of the motor nerve, whereas in others, abortive spikes or localized end-plate potentials were recorded. The proportion of fibers showing very small or no electrical response increased when recordings were made away from the end-plate zone. Reinnervated normal muscle fibers examined 1.5 months after microcauterization of the end-plate region often responded to nerve stimulation with abortive spikes. “Functional denervation” is discussed in terms of presynaptic and postsynaptic defects.

Single smooth muscle cells freshly isolated from the stomach muscularis of the toad Bufo marinus were studied under direct microscopic observation using intracellular microelectrodes. The deviation of the membrane potential from rest was recorded when steps of current were injected into the cell. Outward-going rectification was consistently observed both in the presence of 1.8 mM and higher external concentrations of Ca2+. There was no indication of inward-going rectification even under conditions favoring its demonstration, i.e., when the external concentration of K+ was high (108 mM) and Cl-, low (39.6 mM). In the presence of tetraethylammonium chloride (TEA), there was a marked decrease in the rectification normally observed with depolarizing currents, suggesting that a K+ conductance contributes to the outward-going rectification. This K+ conductance increased by almost two orders of magnitude over the range from -20 to 0 mV, and displayed an e-fold increase with a depolarization as small as 4-7 mV. In response to hyperpolarizing currents, the membrane potential did not always reach a plateau but at times continued to become more negative. The feasibility of the depletion of ions from the caveolae as an explanation for this observation is discussed.

In recent years it has become evident that many cells of nonneuronal origin, including the macrophage, exhibit voltage-dependent conductance previously associated with excitable cells (1, 2). However, because the field of macrophage electrophysiology is relatively new, many questions remain unanswered that pertain to the specific conductances found in these cells and their role in cell function. At present it is known that macrophages exhibit a calcium-activated potassium conductance (3, 4), an inward rectifying potassium conductance (5), and a delayed potassium conductance (6, 7). Spiking behavior, implicating the presence of another voltage-dependent conductance (2), and an electrogenic sodium-potassium pump (8) also have been reported in macrophages. Unfortunately, progress has been hampered by the fact that macrophages are very sensitive to microelectrode-induced damage. Patch clamping substantially reduces penetrationinduced damage and thus has enabled considerable progress to be made in the field of macrophage electrophysiology (1, 4, 6). We have used this technique to confirm and extend some of the observations we previously made using intracellular microelectrodes. This paper reviews the evidence for the presence of an inward rectifying potassium conductance and a calcium- and voltage-dependent potassium conductance in macrophages, and includes some of our more recent data measuring these conductances with patch-clamp techniques.

We describe myotonic electromyographic activity in muscles of genetically dystrophic chickens and show that this activity is antagonized in vivo by doses of phenytoin (DPH) that improve righting ability. To test the possibility that the in vivo effects of DPH can be accounted for by a direct action on skeletal muscle we studied posterior latissimus dorsi fibers in vitro at 23 degrees C with intracellular microelectrodes. Compared to normal fibers, fibers from untreated dystrophic chicks had larger diameters, increased membrane capacitance, longer latencies at rheobase, and a greater tendency to fire repetitively in response to direct stimulation. DPH (10 or 50 micrograms/ml in the bath solution) decreased latencies at rheobase and repetitive firing in fibers from untreated chicks. In DPH-free solution fibers from dystrophic chicks treated chronically with DPH were still abnormal with respect to latencies at rheobase and ease of repetitive firing. The data support the hypothesis that abnormalities of membrane electrical properties are major features of dystrophic chicken muscles and furthermore, show that DPH suppresses, but does not abolish, these abnormalities.

Previous studies indicate that cardiac hypertrophy is associated with altered mechanical properties including prolonged contraction times and mechanical refractory periods and exaggerated aftercontractions in isolated rat papillary muscle preparations. The present study was designed to ascertain whether changes in transmembrane potentials accompany the altered mechanical properties. Responses of papillary muscles from hypertrophied hearts of deoxycorticosterone acetate-treated Wistar Kyoto (WKY) rats were compared to those of nonhypertrophied, sham-treated WKYs. Muscles connected to a force transducer in a bathing chamber containing a 27° oxygenated physiological salt solution were subjected to field stimulation and isometric tension measured. Intracellular microelectrodes were used to measure transmembrane potentials of single impaled cells. When compared to nonhypertrophied controls, the hypertrophied preparations had (1) longer action potentials and contraction times, (2) longer electrical and ...

We used intracellular microelectrodes to study the effect of histamine on oscillatory afterpotentials (OAPs). OAPs were induced by exposing cardiac Purkinje fibers to barium (10(-5) M) and to strophanthidin (3 X 10(-7) M). Histamine consistently increased the amplitude of the barium-induced OAPs. Similar results were obtained in strophanthidin-treated preparations: histamine, in fact, was able to increase the OAPs amplitude and eventually to induce triggered activity. The effect of histamine was antagonized by cimetidine (10(-5) M). The possible role of the OAPs in the arrhythmogenic action of histamine is discussed.

The effects of catecholamines (adrenaline, noradrenaline and isoproterenol) on ionic conductance changes during the generation of action potentials of bullfrog sympathetic and spinal ganglion cells were studied with intracellular microelectrodes. In sympathetic ganglion cells, adrenaline (3X10(-5)-1X10(-3)M) reversibly decreased the peak amplitude and positive after-potential of action potentials, and prolonged the duration of spike potentials without changes in the resting membrane potential and conductance in the Ringer solution. The maximum rates of rise and fall of spike potentials were also decreased. The action of noradrenaline was similar to that of adrenaline, but isoproterenol did not show any effects. Adrenaline (3X10(-5)-3X10(-4)M) markedly depressed the peak amplitude and maximum rate of rise of both TEA-potential and Ca-potential produced either in TEA solution containing TTX or in the isotonic CaCl2 solution. Similar actions were observed with noradrenaline but not isoproterenol. In spinal ganglion cells, catecholamines did not show any effects of the action potentials in Ringer and TEA solutions. It was concluded that adrenaline inhibited the increases in Ca2+, K+ and Na+ conductances during the generation of action potentials of sympathetic ganglion cells.

THERE is considerable indirect evidence for cyclic AMP as the mediator of the chronotropic effects of adrenaline in heart cells. Adrenaline-like changes in electrical activity have been evoked in cardiac preparations by exposure to inhibitors of cyclic AMP phosphodiesterase1,2 or butyryl cyclic AMP derivatives (refs 2 and 3, but compare ref. 4), which are thought to mimic adrenaline because of their presumed ability to raise intracellular cyclic AMP concentrations. This approach has been widely used5 but the interpretation depends heavily on assumptions as to the mode of action of the particular exogenous compound4. We have approached the problem more directly by examining the electrical effects of iontophoretic injection of cyclic AMP from intracellular microelectrodes in cardiac Purkinje fibres.

The sucrose-gap method was adapted to record the action potentials of bullfrog cardiac muscles. The size and configuration of action potentials recorded from atrial or ventricular strips with the sucrose-gap method were similar to those recorded with intracellular microelectrodes. Action potentials could be maintained at a constant amplitude and time-course for up to 3 hours. Furthermore, during recording of action potentials, it was possible to apply DC current to hyperpolarize or depolarize the muscle membrane. Thus the present method in combination with the intracellular microelectrode method will provide valuable information concerning the physiology of cardiac muscle fibers.

Responses to current steps were recorded from pyramidal tract (PT) cells of the cat by means of intracellular microelectrodes. PT cells with resting potentials from -60 to -80 mV set up a well sustained repetitive discharge during stimulation. When comparing fast and slow PT cells, quantitative differences were found between them in the pattern of repetitive firing. Thus, (1) the rheobase is lower in slow PT cells (mean and S.D.; 0.53±0.63 nA) than in fast cells (1.57±1.11 nA). (2) Following stimulation with a current step twice rheobase the latency and the successive interspike intervals are shorter in fast PT cells than in slow cells. (3) The interspike interval distribution shows a greater irregularity in fast PT cells than in slow cells. At firing rates around 30 impulses/sec the coefficient of variation has a mean value of 0.243 for fast PT cells and 0.085 for slow cells. (4) Fast PT cells show a greater decrease of firing rate during the initial 300 msec of current stimulation (adaptation) than do slow cells. The mean value of this initial decrease is 1.85 times the later steady firing rate in fast PT cells and 0.56 times in slow cells. (5) The slope constant of the firing rate-current relationship is larger in fast PT cells, being five times or more than in slow cells. These characteristics of firing pattern are termed “kinetic” and “tonic” for fast and slow PT cells respectively, and their functional meanings are discussed in comparison with other neural organs.

1. Using single intracellular microelectrodes in a bridge circuit, the effect of electrotonus on the activity of motoneurons was explored in excised spinal cords of toads.2. A linear relationship was found between the applied current and the changes of the membrane potential, usually within a range of ±2.5 × 10-9 A, which caused shifts of the membrane potential by ±6 to 8 mV. With stronger currents, the rectifying action of the membrane was usually observed.3. The overshoot of the spike antidromically evoked remained almost constant, either when the membrane was depolarized or hyperpolarized to a certain extent. Stronger such polarizations, however, brought about insufficient or over-compensation.4. The effects of electrotonus on the afterpotentials were studied with similar results to those known from other cells.5. The size of PSP's was reduced by depolarization and augmented within a certain range of hyperpolarization. Monosynaptic EPSP was ineffective in evoking motoneuron spikes in the normal state, but sometimes became effective when the membrane had been suitably depolarized.6. With the combined application of two long square pulses, the motoneurons were excited directly under various background polarizations. Depolarized motoneurons showed responses only with short latency and a rather constant threshold depolarization. On the contrary, hyperpolarized motoneurons responded even with long latencies, the spike arising from a level of depolarization which becomes markedly higher with an increase in the latent time. The accommodation of motoneurons is discussed in connection with these findings.

The Limulus ommatidial potential, as recorded with intracellular microelectrodes, can be divided into three components. These components do not occur in the same portion of the ommatidium. During dark adaptation one can plot the height of the response evoked by a constant stimulus presented at regular intervals of time after the adapting light is removed. Hyperpolarizing current influences the shape of the curve obtained by this means. Increasing potassium ion concentration in the external medium reduces the height of the ommatidial potential. The steady component is decreased more by high potassium than the final component. Removing calcium ion from the medium abolishes the transient component.

WE have previously reported1 that the application of acetylcholine by perfusion or local injection to different central neurones of the mollusc Aplysia depilans, impaled with intracellular microelectrodes, always produced one of the two following reactions: (a) cells of one type, conventionally designated by us as H-neurones, are hyperpolarized and inhibited by the drug (Fig. 1, H1), and (b) cells of another type, or D-neurones, are depolarized and excited (Fig. 1, D1).

1. Intracellular microelectrodes were used to record spontaneous and evoked inhibitory junction potentials (IJP) from the circular muscle layer of the mid-distal region of the mouse isolated colon in the presence of nifedipine (1 micromol/L) and hyoscine (1 micromol/L). 2. The length of the tissue preparation (> 1 cm) or the presence of the mucosa had no effect on the frequency of spontaneous IJP. 3. Hexamethonium (500 micromol/L) reduced the frequency of spontaneous IJP to approximately 70% of the control frequency, whereas D-tubocurarine (280 micromol/L) reduced the frequency to approximately 17% of control. Apamin (250 nmol/L) abolished all spontaneous IJP activity. 4. The greater inhibition of spontaneous IJP in the presence of D-tubocurarine compared with hexamethonium is discussed as a possible 'apamin-like' effect. 5. Although electrically evoked IJP (single pulse at 15 V, 0.6 msec) were not significantly affected by hexamethonium, D-tubocurarine and apamin reduced the amplitude of evoked IJP to approximately 65 and 50% of control, respectively. 6. These results suggest that the properties of spontaneous IJP cannot be inferred by a study of evoked IJP alone.

Aldosterone plays a major role in atrial structural and electrical remodeling, in particular through Ca2+-transient perturbations and shortening of the action potential. The Ca2+-activated non-selective cation channel Transient Receptor Potential Melastatin 4 (TRPM4) participates in atrial action potential. The aim of our study was to elucidate the interactions between aldosterone and TRPM4 in atrial remodeling and arrhythmias susceptibility. Hyperaldosteronemia, combined with a high salt diet, was induced in mice by subcutaneously implanted osmotic pumps during 4 weeks, delivering aldosterone or physiological serum for control animals. The experiments were conducted in wild type animals (Trpm4+/+) as well as Trpm4 knock-out animals (Trpm4-/-). The atrial diameter measured by echocardiography was higher in Trpm4-/- compared to Trpm4+/+ animals, and hyperaldosteronemia-salt produced a dilatation in both groups. Action potentials duration and triggered arrhythmias were measured using intracellular microelectrodes on the isolated left atrium. Hyperaldosteronemia-salt prolong action potential in Trpm4-/- mice but had no effect on Trpm4+/+ mice. In the control group (no aldosterone-salt treatment), no triggered arrythmias were recorded in Trpm4+/+ mice, but a high level was detected in Trpm4-/- mice. Hyperaldosteronemia-salt enhanced the occurrence of arrhythmias (early as well as delayed-afterdepolarization) in Trpm4+/+ mice but decreased it in Trpm4-/- animals. Atrial connexin43 immunolabelling indicated their disorganization at the intercalated disks and a redistribution at the lateral side induced by hyperaldosteronemia-salt but also by Trpm4 disruption. In addition, hyperaldosteronemia-salt produced pronounced atrial endothelial thickening in both groups. Altogether, our results indicated that hyperaldosteronemia-salt and TRPM4 participate in atrial electrical and structural remodeling. It appears that TRPM4 is involved in aldosterone-induced atrial action potential shortening. In addition, TRPM4 may promote aldosterone-induced atrial arrhythmias, however, the underlying mechanisms remain to be explored.

Spontaneous junctional potentials (SJP) in guinea-pig vas deferens were investigated with intracellular microelectrodes. In some of the cells examined, SJP was atropine-sensitive and presumably cholinergic in nature. This finding stands in contrast to the generally accepted concept that all of SJPs in vas deferens are atropine-resistant and noradrenergic in nature. It seems more likely from this result that there exist two types of SJPs in vas deferens; one is noradrenergic type and the other is cholinergic type.