Intrinsic burst generation of preinspiratory neurons in the medulla of brainstem-spinal cord preparations isolated from newborn rats

In brainstem-spinal cord preparations isolated from newborn rats, intrinsic burst-generating properties of preinspiratory (Pre-I) neurons in the rostral ventrolateral medulla, which have been suggested to be primary respiratory rhythm-generating neurons, were studied by perforated whole-cell recordings using the antibiotic nystatin. Nystatin causes small pores to be formed in the cells, through which pass small monovalent ions. For blockade of chemical synaptic transmission, perfusate Ca²⁺ concentration was lowered to 0.2 mM and the Mg²⁺ concentration was increased to 5 mM. In Iow-Ca²⁺, high-Mg²⁺ solution (referred to here as low Ca), 10 of 55 Pre-I neurons generated rhythmic bursts (burst type), 14 fired tonically (tonic type), and 31 were silent (silent type). Burst-type neurons showed periodic depolarization of 5–12 mV in low Ca, at a rate of 12±6.5/min. Hyperpolarization of the membrane caused decrease in or disappearance of the periodic depolarization and prolongation of the cycle period. Thus, the burst generations were voltage dependent. The firing frequency of tonictype neurons was 2.3±1.6 Hz and was decreased by hyperpolarization. In 6 of these neurons, the firing patterns changed to burst patterns during continuous hyperpolarization. Membrane depolarization by continuous outward current injection into some silent-type neurons (3 of 11 tested) induced bursting activity. Activity of C4 and Pre-I neurons was completely silent with 0.1–1 M tetrodotoxin (TTX) added to the standard perfusate. In low Ca, burst-type neurons (n=3) were also silent with 1 M TTX perfusion. Inspiratory neurons either became silent (n=4) or fired tonically (n=1) in low Ca. The present study by perforated whole-cell recordings confirmed that some Pre-I neurons possess intrinsic burst-generating properties, which were not attributable to phasic synaptic inputs.     

Depolarizing IPSPs and depolarization by GABA of rat neostriatum cells in vitro

Summary In neostriatal slices pretreated with sodium pentobarbital (100 M) and 4-aminopyridine (50 M), intrastriatal stimulation elicited EPSPs followed by a slowly decaying depolarization which lasted about 200 ms and was associated with a membrane conductance increase and a suppression of spike potentials. This depolarizing inhibitory synaptic action could be blocked by picrotoxin (50 M) or bicuculline (50 M). The reversal potential for the slowly decaying depolarization was –57 to –62 mV, i.e. it was positive with respect to the resting membrane potential (¯x = –67 mV). GABA, injected into the tissue in the vicinity of the recording electrode by pressure application, or added to the perfusate (10 M–1 mM), depolarized the cells and reduced both the membrane resistance and the amplitude of EPSPs. The reversal potential of GABA depolarization was found in a potential range approximating that of the slowly decaying depolarization. These results are compatible with the assumption that GABA is the transmitter of an intrinsic inhibition in rat neostriatum, but indicate that GABA-mediated IPSPs of neostriatal cells in vitro are depolarizing at the resting membrane potential. The possible reasons for this are discussed.     

Adenosine enhances afterhyperpolarization and accommodation in hippocampal pyramidal cells

Adenosine added to the perfusion fluid of rat hippocampal slices at 10 mol · l^–1 enhanced long lasting afterhyperpolarizations after single action potentials, bursts of action potentials or calcium spikes. Accommodation of firing during a depolarizing pulse was potentiated. An increase in calcium dependent potassium conductance is likely to mediate these effects. Adenosine at 50 mol·l^–1 induced a hyperpolarization accompanied by a reduction in input resistance. The hyperpolarization could be reversed at –85 mV. In TTX and TTX-barium treated slices the amplitude of the slow spike was decreased. This may result from a shunting of inward current in the dendrites due to an adenosine induced increase in potassium conductance. It is suggested that adenosine reduces pre- and postsynaptic exicatory signals principally by enhancing one or more potassium conductances. This effect is a powerful means for modulation of neuronal excitability and synaptic efficacy and can explain the antiepileptic activity of adenosine.     

Inward rectifier and low-threshold calcium currents contribute to the spontaneous firing mechanism in neurons of the rat suprachiasmatic nucleus

Intracellular and voltage-clamp studies were carried out to clarify the mechanism for spontaneous firing activity in neurons of the suprachiasmatic nucleus (SCN) of rat hypothalamic brain slices in vitro. SCN neurons displayed spontaneously firing action potentials that were preceded by a depolarizing pre-potential and followed by a short spike after-hyperpolarization (AHP). Injection of inward current with a duration longer than 50 ms resulted in a depolarizing voltage sag on hyperpolarizing electrotonic potentials. The inward rectification was depressed by bath application of caesium (1 mM) but not by barium (500 M). SCN neurons also showed a rebound depolarization associated with spike discharge (anodal break) in response to relaxation of hyper polarizing current injection. The rebound depolarization was reduced by nominally zero calcium. Cadmium (500 M), cobalt (1 mM) or caesium (1 mM) but not nicardipine also depressed the rebound depolarization. Under voltage-clamp conditions, hyperpolarizing steps to membrane potentials negative to approximately –60 mV caused an inward rectifier current, probably H current (I _H), which showed no inactivation with time. Bath application of caesium (1–2 mM) suppressed I _H. Caesium (2 mM) depressed the slope of the depolarizing spike pre-potential, resulting in a prolongation of the interspike interval of tonic firing neurons. We conclude that both the inward rectifier current, I _H, and the low-threshold calcium current contribute to the spike prepotential of spontaneous action potentials in firing neurons of the rat SCN.     

Spike trains and signaling modes of neurons in the ferret lateral geniculate nucleus

Intracellular recordings were used to examine the action potential firing modes of cells in the ferret lateral geniculate nucleus (LGN) in vitro. We compared the effects of altered membrane voltage on patterns of action potential trains evoked by direct current injection and by retinal afferent (synaptic) stimulation. The results confirm that LGN cells in the ferret can fire action potentials in the burst and tonic modes that have been described previously for other species. At depolarized membrane potentials, LGN neurons respond to sustained depolarization with short-latency trains of action potentials whose frequency is directly proportional to the amount of current injected. At hyperpolarized membrane potentials, LGN cells enter burst mode, in which depolarizing inputs are differentiated into brief high-frequency discharges whose latency varies with membrane potential. We also observed a mixed mode, in which LGN cell responses to synaptic or injected currents within a narrow range of membrane potentials reflect aspects of both burst and tonic firing simultaneously. Thus a striking consequence of the interplay among voltage-dependent membrane conductances in thalamic cells is wide variability in length, duration, and latency of spike discharges elicited by identical stimuli. These results also suggest that the concept that LGN cells display only two active response modes must be expanded to include varying amounts of delay and the possibility of mixed discharges.     

β-Adrenoceptor-mediated depolarization of the resting membrane in guinea-pig papillary muscles: changes in intracellular Na+, K+ and Cl− activities

Effects of -adrenergc stimulation on the membrane potential and intracellular Na⁺, K⁺ and Cl^– activities were examined in isolated guinea-pig ventricular muscles using conventional and ion-selective elctrodes. Isoproterenol in concentrations of 30 nM–1 M produced a transient depolarization followed by a slight hyperpolarization in electrically stimulated or quiescent papillary muscles. The negative logarithm of the concentration producing 50% maximum effect (pD₂) for the membrane-depolarizing effect of isoproterenol was smaller than that for the positive inotropic effect, suggesting that a higher level of cAMP accumulation is required to produce the transient depolarization. Whereas the isoproterenol(1 M)-induced depolarization was not blocked by tetrodotoxin (10 M), nifedipine (10 M), Cs⁺ (5 mM), Ba²⁺ (0.3 mM), amiloride (1 mM) or ouabain (10 M), it was significantly attenuated by anthracene-9-carboxylic acid (1 mM), a Cl^–-channel blocker. Intracellular K⁺ activity increased, whereas intracellular Na⁺ activity slightly decreased during the transient depolarization. Intracellular Cl^– activity significantly decreased during the isoproterenol-induced depolarization of the resting membrane. These results suggest that an inward current resulting from outward Cl^– movement, rather than inward Na⁺ movement, may be involved in the -adrenoceptor-mediated membrane depolarization.     

Actions of glutamic acid on spinal neurones

Summary Spinal neurones from the 6th and 7th lumbar segments of cats were recorded intracellularly. Glutamic acid (GLUT) was applied extracellularly by means of the microelectrophoretic technique from another parallel electrode and caused, in almost all cells, a depolarization of the cell in association with conductance change. In some cases, initial depolarization occurred with no detectable conductance change.Motoneurones could not be brought to continous firing except in two cases. High doses of GLUT (up to 2000 nA) caused maximal depolarization up to-30 mV (range -32 mV to -24 mV). The soma conductance was increased at this time by about 75% (range 60–90%).The spikes and both EPSPs and IPSPs were attenuated by shunting. The peak latency of the EPSP was shortened. Spikes evoked by different EPSPs were found to exhibit different sensitivity to GLUT. Ocassionally, the IPSPs were initially increased following depolarization. The IPSP was sometimes reversed after the termination of GLUT application. The after-hyperpolarization following the antidromic spike of some motoneurones was converted into an after-depolarization for 1–2 min.The present data show that the following criteria for an excitatory transmitter can be met by GLUT; (1) strong depolarizing action presumably associated with Na⁺ influx, (2) an associated conductance change and (3) an equilibrium potential. It could not be proven that the EPSP reversal point and the equilibrium potential for GLUT are the same.Fellow of the Medical Research Council of Canada.     

Acceleration of relaxation by hyperpolarization of the crayfish muscle fibre membrane

Summary The membrane of single crayfish muscle fibres was depolarized to –50 to zero mV for 50 or 500 ms by a voltage clamp through two microelectrodes. Simultaneously, clamp current, isometric force development and its first time derivative were recorded. After the depolarization pulse, the membrane potential was either clamped back to the holding potential of about –70 mV (repolarization), or clamped to a more negative potential of –95 to –125 mV (hyperpolarization). During such hyperpolarizations, relaxations following large contractions were accelerated. If e. g. the contractions were triggered by depolarizations to –29 to zero mV of 500 ms duration, the average increase in the maximum rate of relaxation due to hyperpolarization was 27±3% (s.e.). The acceleration of relaxation by hyperpolarization started about 100 ms after the end of the depolarization pulse. This latent period was observed after depolarization pulses of 50 as well as 500 ms duration. If chloride in the bathing solution was replaced by nitrate, the acceleration of relaxation by hyperpolarization was increased. If chloride was replaced by propionate, relaxation was slowed. The acceleration of relaxation by hyperpolarization was smaller in propionate than in chloride saline. The latent period for acceleration to appear after the end of the depolarization pulse increased to 450 ms in propionate saline.—The results seem to indicate a state of excitation in parts of the transverse tubular system triggered by depolarization, which cannot fully be terminated by a repolarization of the external membrane for several 100 ms and thus slows relaxation.This study was supported by the Deutsche Forschungsgemeinschaft.     

Voltage-dependent sodium and potassium,but no calcium conductances in DDT1 MF-2 smooth muscle cells

Voltage-dependent inward and outward membrane currents were investigated in the DDT₁ MF-2 smooth muscle cell line using the whole-cell patch-clamp technique. Application of a pulse protocol with subsequent depolarizing voltage steps elicited an inactivating inward current and a non-inactivating outward current. The outward current was activated at membrane potentials more positive than –35 mV, with _act=30–40 ms. The outward current was blocked by tetraethylammonium (NEt₄Cl) and 3, 4-aminopyridine in a dose-dependent manner (EC₅₀ of 5 mM and 0.5 mM, respectively). The amplitude of the outward current was linked to the potassium equilibrium potential (V_ek), and tail currents reversed near V_ek. The outward current was completely abolished when intracellular potassium was substituted by 106 mM caesium and 20 mM NEt₄Cl. The inward current was activated at potentials more positive than –30 mV with _act of 1.6–2.5 ms, and with _inact of 1.7–3.0 ms. Steady-state inactivation was 50% at a holding potential of –40 mV. The inward current was blocked by tetrodotoxin (EC₅₀ of 0.15 M) and dependent on the reversal potential for sodium. Voltage-dependent calcium currents could not be detected. Further, the cytoplasmic free calcium concentration, as measured using Indo-1 fluorescence, was not changed during high-potassium(40 mM)-induced depolarization. In contrast, contraction of freshly obtained hamster vas deferens tissue elicited by high-potassium(40 mM)-induced depolarization was largely inhibited by diltiazem (20 M). These findings showed that voltage-dependent calcium channels are not functional in DDT₁ MF-2 smooth muscle cells in contrast to freshly obtained Syrian hamster vas deferens smooth muscle. It is concluded that at least two populations of voltage-dependent channels are present in DDT₁ MF-2 smooth muscle cells, conducting a slow outward rectifying current carried by potassium, and a fast, inactivating sodium current.     

Functional expression and properties of the human skeletal muscle sodium channel

Full-length deoxyribonucleic acid, complementary (cDNA) constructs encoding the-subunit of the adult human skeletal muscle Na⁺ channel, hSkM1, were prepared. Functional expression was studied by electrophysiological recordings from cRNA-injectedXenopus oocytes and from transiently transfected tsA201 cells. The Na⁺ currents of hSkM1 had abnormally slow inactivation kinetics in oocytes, but relatively normal kinetics when expressed in the mammalian cell line. The inactivation kinetics of Na⁺ currents in oocytes, during a depolarization, were fitted by a weighted sum of two decaying exponentials. The time constant of the fast component was comparable to that of the single component observed in mammalian cells. The block of hSkM1 Na⁺ currents by the extracellular toxins tetrodotoxin (TTX) and -conotoxin (CTX) was measured. The IC₅₀ values were 25 nM (TTX) and 1.2 M (CTX) in oocytes. The potency of TTX is similar to that observed for the rat homolog rSkM1, but the potency of CTX is 22-fold lower in hSkM1, primarily due to a higher rate of toxin dissociation in hSkM1. Single-channel recordings were obtained from outside-out patches of oocytes expressing hSkM1. The single-channel conductance, 24.9 pS, is similar to that observed for rSkM1 expressed in oocytes.     

Intracellular analysis of potentiation of CA1 hippocampal synaptic transmission by phorbol ester application

Summary (1) The effect of active and inactive phorbol esters on synaptic transmission and on membrane properties of CA1 pyramidal cells in hippocampus have been analyzed by intracellular recording. (2) 4-phorbol-12,13 dibutyrate (PDBu), but not the -isomer, increased the firing probability, reduced the spike latency and enhanced the EPSP amplitude in response to synaptic activation. The effect was similar to the changes seen in long term potentiation. After PDBu addition it was possible to elicit further enhancement by tetanization, but not after PDBu administration. (3) A slowly developing hyperpolarization was seen after active phorbol ester application without apparent changes in the soma input resistance. (4) Active phorbol esters reduced the slow afterhyperpolarization (AHP) in these cells without affecting the intermediate AHP.     

Disinhibition of hippocampal pyramidal cells during the transition into theta rhythm

The activity of hippocampal complex-spike cells (presumed pyramidal cells) and theta cells (presumed interneurons) was examined during transitions from non-theta electroencephalogram (EEG) states to theta EEG states in freely moving and sleeping rats. Theta cell firing rates were significantly depressed in a 1-s period centered on the EEG transition relative to the surrounding 1-s periods (normalized rates±SEM): 1.05±0.02 for the non-theta period, 0.59±0.03 for the transition period, and 1.36±0.04 for the theta period (n = 26 cells). Conversely, complex-spike cell firing was significantly increased during the transition period: 0.51±0.11 for the non-theta period, 2.24±0.19 for the transition period, and 0.24±0.04 for the theta period (n = 27 cells). This diametrically altered activity indicates that theta cells must be actively inhibited during the transition. The increased activity in complex-spike cells during the transition may be simply a release from inhibitory control by interneurons. The pattern of theta cell inhibition together with increased complex-spike cell activity appears to be a general property of transitions into the theta EEG state, irrespective of behavior. It is suggested that increased activity in septal afferents (GABAergic cell activity greater than cholinergic cell activity) initially inhibits hippocampal interneurons. The inhibition is not sustained because of an activity-dependent decrease in the potency of the septointerneuronal inhibition, leaving the rhythmic excitatory (cholinergic) septointerneuronal inputs, together with principal cell inputs, to increase interneuron firing rates.     

Prejunctional effects of anticholinesterase drugs at the endplate

Anticholinesterase drugs induce antidromic firing of motor axons in mammalian nerve-muscle preparations. Antidromic firing is observed in response to a conditioning stimulus applied to the nerve and hence is known as back-firing; the present investigation intends to clarify the mechanisms underlying this phenomenon. Acetylcholinesterase (AChE) was inhibited with neostigmine. Antidromic action potentials were recorded extracellularly from the nerve trunk. When the muscle was cut on one side of the main endplate region 0.1–0.2 cm away from the outermost myelinated nerve branches, back-firing gradually disappeared. This indicated that postsynaptic factors at least contribute to the generation of back-firing. One can conceive of a plausible mechanism whereby postsynaptic events influence presynaptic excitability: potassium ions leaking from extensively depolarized muscle fibers (block of AchE) might cause depolarization of nerve endings at some endplates (potassium hypothesis). Postsynaptic potassium currents were calculated according to a simple mathematical model of the muscle membrane. These simulations predict high postsynaptic potassium currents under the following experimental conditions: (1) low concentration of extracellular potassium, (2) exchange of extracellular chloride with nitrate, (3) small fiber diameter. Back-firing was found to increase under such conditions. The notion that postsynaptic potassium efflux may influence presynaptic membrane polarization was further substantiated: when carbamoylcholine chloride (Carbachol; 10 mol · 1^–1) was added to the bathing fluid, miniature endplate current frequency could be increased at some voltage clamped endplates by depolarizing the muscle fiber well below the potassium equilibrium potential.This work was supported by the Deutsche Forschungsgemeinschaft, SFB 38, project N     

Extracellular K+ accumulations and synchronous GABA-mediated potentials evoked by 4-aminopyridine in the adult rat hippocampus

Transient changes in extracellular potassium concentration ([K⁺]₀) and field potentials were evoked by 4-aminopyridine (4-AP; 50–100 M) and recorded with ion-selective microelectrodes in CA1b, CA3b and dentate sectors of adult rat hippocampal slices. Long-lasting field potentials recurred at a frequency of 1/60 s (0.016±0.003 Hz) in association with increases in [K⁺]₀ which were largest and most sustained in the dendritic regions where afferent fibers terminate (dentate>CAl>CA3) and in the hilus. In stratum radiatum of CA1 or stratum moleculare of the dentate these fields had a peak amplitude of 1.4±0.29 mV, duration 8.3±1.6 s, and were accompanied by increases in [K⁺]₀ of 1.8±0.22 mM that lasted 32±5.5 s (n = 17 slices). Interictal epileptiform potentials, which were brief (<0.2 s) and more frequent at 1/3 s (0.30±0.02 Hz) were also present in CA1, CA3 and the hilus and associated with small increases in [K⁺]₀ (0.5 mM, duration 2 s). Interictal activity was blocked by 6-cyano-7-nitroquinoxalone-2,3-dione (CNQX; 5–20 M); the slow, less frequent potentials were resistant to both CNQX and dl-2amino-5-phosphonovaleric acid (APV; 50 M) and reversibly blocked (or attenuated by 80%) by bicuculline methiodide (BMI) (25–100 M). The BMI-sensitive potentials were also abolished by baclofen (100 M), an effect which was reversed by 2-OH-saclofen (100 M). Focal application of KCl or GABA in the absence of 4-AP evoked long-lasting field and [K⁺]₀ potentials which were similar to those evoked by 4-AP but more sustained. The proportional relationship between the amplitudes of field and K⁺ potentials with GABA closely resembled that observed for 4-AP; in contrast the slope of KCl-evoked responses was lower. Our results demonstrate that in the adult rat hippocampus 4-AP induces in many different regions accumulations of [K⁺]₀ in synchrony with the long-lasting field potentials, which are known to correspond to an intracellular long-lasting depolarization of the pyramidal cells. These changes are smaller than those which occur in the immature rat hippocampus — which may be related to differences in Na-K-ATPase and susceptibility to seizures. These events involve the activation of GABA_A receptors, are under the modulatory control of GABA_B receptors, and likely arise from the activity of GABAergic interneurons and/or afferent terminals. The long-lasting field potentials appear to reflect mainly the direct depolarizing actions of GABA and to a much more limited extent the associated accumulation of [K⁺]₀.     

Electrophysiological and repetitive firing properties of neurons in the superficial/middle layers of the human neocortex maintained in vitro

Conventional intracellular recordings were made from neurons located in the superficial/middle layers of human temporal neocortical slices obtained from patients undergoing neurosurgical procedures for the treatment of epilepsy or brain tumour. In most of the neurons, inward membrane rectification was observed when the cell was depolarized or hyperpolarized from rest by intracellular injection of positive or negative current pulses. Bath application of tetrodotoxin abolished the depolarizing inward rectification, but not the anomalous rectification in the hyperpolarizing direction. Single action potential firing was followed by a fast afterhyperpolarization, a depolarizing afterpotential and a medium afterhyperpolarization, while a slower afterhyperpolarization was seen following repetitive firing. Blockade of Ca² channels with Cd² diminished all three types of afterhyperpolarization. Although the repetitive firing pattern in all cells indicated that they discharge in a regular-spiking fashion, 63% of the cells fired tonically in the initial part of discharge, while the remaining 37% of the cells fired phasically. Frequencycurrent plot for the initial interspike intervals during long depolarizing pulses revealed primary and secondary ranges of firing. Spike frequency adaptation was also observed. In conclusion, our experiments indicate that human neocortical cells in the superficial/middle layers display electrophysiological characteristics that are similar to those described in rodent and feline neocortices.     

Nociceptive neurones in the superficial dorsal horn of cat lumbar spinal cord and their primary afferent inputs

Summary The morphology, background activity and responses to stimulation of primary afferent inputs of small neurones in the superficial dorsal horn which could only be excited from the skin by noxious stimulation were investigated by intracellular recording and ionophoresis of HRP. Neurones which gave similar responses to afferent stimulation were morphologically heterogeneous with respect to dendritic tree geometry and axonal projection, but were located around the lamina I/II border. Cutaneous excitatory receptive fields responding to noxious stimulation were generally small; most neurones had more extensive inhibitory fields responding to innocuous mechanical stimulation, in many cases overlapping the excitatory fields. Generally, stimulation of the excitatory field resulted in depolarization of the neurone and increased action potential firing, and stimulation of the inhibitory field resulted in hyperpolarization. Electrical stimulation of peripheral nerves revealed the existence of converging excitatory inputs carried by different fibre groups, and all neurones received an inhibitory input activated at low threshold. Excitatory responses were short-lived and occurred consistently in response to repeated stimulation. Central delay measurements gave evidence of a number of A monosynaptic inputs but only one A monosynaptic input; inhibitory inputs along A fibres were polysynaptic. The constant latency and regularity of the C response suggested monosynaptic connections. Low intensity stimulation of inhibitory inputs elicited a short-lived i.p.s.p. which increased in amplitude with increasing stimulus strength until it disappeared into a more prolonged hyperpolarization. This was associated with inhibition of background action potentials, and increased in duration with increasing stimulus strength up to C levels, indicating an A and C component. It is suggested that the level of excitability of these neurones depends on the relative amounts of concurrent noxious and innocuous stimulation, and that the resultant output, which is conveyed mainly to other neurones within the spinal cord, could modulate reflex action at the spinal level as well as affecting components of ascending sensory pathways.Supported by grant no. 11853/1.5 from the Wellcome Trust     

Mechanisms of Hyperpolarizing Effect of GABA on Resting Potential of the <Emphasis Type="Italic">Lumbricus Terrestris</Emphasis> Muscular Wall Somatic Cells

GABA, baclofen, isoguvacine increase, and cis-4-aminocrotonic acid does not modify resting membrane potential of muscle cells. Bicuculline, phaclofen, N-ethylmaleimide, chlorpromazine, verapamil, and removal of Ca²⁺ from bathing solution abolished the effect of baclofen, while U73122 and D609 were ineffective in this respect. The authors conclude that the Lumbricus terrestris muscle cells contain GABAergic structures similar to a- and b-receptors. Activation of GABA receptors induced Cl^- inward current and Ca²⁺ entry with subsequent activation of calmodulin-like proteins, which causes membrane hyperpolarization by increasing the effect of pumping potential on resting membrane potential.__________Translated from Byulleten Eksperimentalnoi Biologii i Meditsiny, Vol. 139, No. 2, pp. 219–222, February, 2005     

Repetitive impulses generated in fast and slow pyramidal tract cells by intracellularly applied current steps

Summary 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.     

Forskolin-induced block of delayed rectifying K+ channels in pancreaticβ-cells is not mediated by cAMP

K⁺ channels in the membrane of murine pancreatic -cells were studied using the patch-clamp technique. The delayed outward current was activated in whole-cell experiments by depolarizing voltage pulses to potentials between –30 mV and 0 mV. Forskolin blocked the current rapidly (<5 s) and reversibly with 50% inhibition at 13 M. The inhibition did not depend on a stimulation of the adenylate cyclase since it occurred even in presence of 1 mM cAMP in the pipette solution which replaced the cytoplasm. Membrane permeant cAMP analogues and phosphodiesterase inhibitors did not influence the delayed outward current. In experiments on outside-out patches forskolin (100 M) shortened the openings of a channel of about 10 pS conductance at 0 mV and a time course of activation and inactivation similar to the whole-cell current. Another smaller, slowly activating channel and the Ca²⁺- and ATP-dependent K⁺ channels were influenced only weakly or not at all. It is therefore concluded that the 10-pS channel generates most of the delayed outward K⁺ current in murine pancreatic -cells. The Ca²⁺-independent part of the delayed outward current in bovine adrenal chromaffin cells was also blocked by forskolin (100 M).     

Processing vibratory stimuli in isolated frog muscle spindle

Summary Experiments were performed on isolated frog muscle spindle receptors to study the particular transducer and encoder mechanisms involved in the signal transfer of high frequency sinusoids (vibration). In order to systematically investigate the signal transfer over the entire dynamic range of the receptor, vibration stimuli were applied to the intrafusal muscle bundle at different prestretch levels, so that the isolated receptor potential or the afferent impulse train were recorded at different operating points. The vibration-induced receptor potential displayed severe distortion, because the depolarization during stretch rose steeply, whereas the repolarization transient during release of stretch declined more slowly. The positive peak velocity values of the depolarization transient increased with increasing stimulus frequency, although the ac-component of the receptor potential decreased. The negative peak velocity values of the repolarization transient remained constant throughout the frequency range. The amplitude of the receptor potential grew larger when vibration of constant amplitude was applied at increasing levels of prestretch, revealing another non-linearity of the transducer. These two types of non-linearity were influential in determining the afferent discharge pattern. Each fast depolarization transient facilitated the generation of a single action potential, which therefore could be firmly phase-locked to a small segment of the vibratory movement. Due to its short risetime, the depolarization transient tended to prevent multiple firing during one stimulus cycle. The prolonged depolarizing afterpotential of the evoked action potential operated in the same direction. Increasing prestretch greatly enhanced the responsiveness of the spindle to vibration. Thus, under appropriate conditions, the afferent discharge was driven in 11 synchrony with the vibration. An analysis is given of the after-effects of repetitive activity at the receptor site. The progressive decline of the mean membrane voltage during long lasting stimulation and the post-tetanic hyperpolarization (off-effect) on termination of the vibration suggest the action of an electrogenic pumping mechanism. As a consequence, the afferent impulse train possessed a complex structure segmented into several transient and steady states, which differed in impulse rate, phase response, and in the degree of phaselocking