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H A Lester

Publications and source records attributed to H A Lester.

196 records · Page 11Linked to original sources
PubMed1978

Role of voltage-sensitive receptors in nicotinic transmission.

This paper compares the conductance induced by bath-applied acetyl-choline (ACh) and by the same transmitter released from nerve terminals at Electrophorus electroplaques. For the former case, dose-response relations are characterized by the maximal agonist-induced conductance, rgamma (130 mmho/cm2), and by the concentration which induces half this conductance; this concentration is termed Kapp and equals 50 micron at -85 mV. For the latter case, neurally evoked postsynaptic currents (PSCs) are characterized by the peak conductance during strongly facilitated release, gPSC, and by the rate constant for decay, alpha. Since gPSC roughly equals rgamma, it is concluded that the PSC activates nearly all available receptor channels. These and other data agree with recent estimates that during the growth phase of the quantal response, (a) the ACh concentration is at least several hundred micromolar; and (b) most nearby channels are activated. However both alpha and Kapp increase during depolarization, at a rate of about e-fold per 86 mV. These observations on voltage sensitivity suggest that a suprathreshold synaptic event is rapidly terminated because the action potential abruptly releases ACh molecules from receptors.

Acetylcholine↗
PubMed1978

Analysis of sodium and potassium redistribution during sustained permeability increases at the innervated face of Electrophorus electroplaques.

Cholinergic agonists cause an increase in the membrane permeability of Na and K at the innervated face of Electrophorus electroplaques. Therefore, sustained exposure to agonist reduces Na and K concentration gradients. There gradients are monitored with voltage-clamp sequences and pharmacological treatments that selectively measure the Nernst potentials for individual ions. EK is normally near--90 mV but moves toward zero during bath application of agonist. Depolarizations by bath-applied agonist measure primarily this shift of EK, not short-circuiting of EK by the agonist-induced conductance. After a rapid jump of agonist concentration, there is a fast (millisecond) depolarization due to the conductance increase, followed by a much slower additional "creep" due to the shift in EK. Sodium replaces the lost intracellular potassium: ENa, normally very positive, also moves toward zero. The shifts in EK and ENa are normally reversible but become permanent after blockade of the Na-K pump. In the presence of agonist, the shifts can be driven further by passing current of the appropriate polarity. Similar ion redistribution occurs with other drugs, such as batrachotoxin and nystatin, which induce prolonged increases in Na permeability. The redistributions cause little net change in the reversal potential of the neurally evoked postsynaptic current.

Animals↗
PubMed1977

Rates and equilibria at the acetylcholine receptor of Electrophorus electroplaques: a study of neurally evoked postsynaptic currents and of voltage-jump relaxations.

Kinetic measurements are employed to reconstruct the steady-state activation of acetylcholine [Ach] receptor channels in electrophorus electroplaques. Neurally evoked postsynaptic currents (PSCs) decay exponentially; at 15 degrees C the rate constant, alpha, equals 1.2 ms(-1) at 0 mV and decreases e-fold for every 86 mV as the membrane voltage is made more negative. Voltage-jump relaxations have been measured with bath-applied ACh, decamethonium, carbachol, or suberylcholine. We interpret the reciprocal relaxation time 1/tau as the sum of the rate constant alpha for channel closing and a first-order rate constant for channel opening. Where measureable, the opening rate increases linearly with [agonist] and does not vary with voltage. The voltage sensitivity of small steady-state conductances (e- fold for 86 mV) equals that of the closing rate alpha, confirming that the opening rate has little or no additional voltage sensitivity. Exposure to alpha-bungarotoxin irreversibly decreases the agonist-induced conductance but does not affect the relaxation kinetics. Tubocurarine reversibly reduces both the conductance and the opening rate. In the simultaneous presence of two agonist species, voltage-jump relaxations have at least two exponential components. The data are fit by a model in which (a) the channel opens as the receptor binds the second in a sequence of two agonist molecules, with a forward rate constant to 10(7) to 2x10(8) M(-1)s(-1); and (b) the channel then closes as either agonist molecule dissociates, with a voltage-dependent rate constant of 10(2) to 3x10(3)s(-1).

Acetylcholine↗
PubMed1975

Relaxation measurements on the acetylcholine receptor.

In Electrophorus electroplaques, the agonist-induced postsynaptic conductance depends on membrane potential. During steady exposure to agonists, after a voltage step the conductance relaxes on a millisecond time scale, exponentially approaching a new equilibrium value. The relaxation rate constant k is an instantaneous function of voltage, insensitive to the past or present conductance. Two components sum to form k. A concentration-sensitive component increases linearly with agonist concentration and decreases during desensitization or exposure to curare. Thus this component reflects the average frequency at which acetylcholine receptors are opening. The voltage-sensitive component, obtained by extrapolating k to zero agonist concentration, increases at more positive potentials. For acetylcholine, the voltage-sensitive component equals the rate constant for the exponential decay of postsynaptic currents; it thus seems to be the closing rate for active receptors. The voltage-sensitive component has the relative amplitudes acetylcholine less than carbamoylcholine less than decamethonium, and for each agonist equals the closing rate determined from "noise" measurements at neuromuscular junctions. The kinetic data explain several aspects of the steady-state conductance induced by agonists, but shed no light on apparent cooperative effects.

Acetylcholine↗
PubMed1975

Conductance increases produced by bath application of cholinergic agonists to Electrophorus electroplaques.

When solutions containing agonists are applied to the innervated face of an Electrophorus electroplaque, the membrane's conductance increases. The agonist-induced conductance is increased at more negative membrane potentials. The "instantaneous" current-voltage curve for agonist-induced currents is linear and shows a reversal potential near zero mV; chord conductances, calculated on the basis of this reversal potential, change epsilon-fold for every 62-mV change in potential when the conductance is small. Conductance depends non-linearly on small agonist concentrations; at all potentials, the dose-response curve has a Hill coefficient of 1.45 for decamethonium (Deca) and 1.90 for carbamylcholine (Carb). With agonist concentrations greater than 10(-4) M Carb or 10(-%) M Deca, the conductance rises to a peak 0.5-1.5 min after introduction of agonist, then declines with time; this effect resembles the "desensitization" reported for myoneural junctions. Elapid alpha-toxin, tubocurarine, and desensitization reduce the conductance without changing the effects of potential; the apparent dissociation constant for tubocurarine is 2 X 10(-7) M. By contrast, procaine effects a greater fractional inhibition of the conductance at high negative potentials.

Animals↗
PubMedDate not supplied

New photoactivatable cyclic nucleotides produce intracellular jumps in cyclic AMP and cyclic GMP concentrations.

The cyclic nucleotides cyclic AMP and cyclic GMP are important intracellular messengers mediating the responses to neurotransmitters and neurohormones and regulating cellular function over a wide range of time scales. Despite the widespread acceptance of this second messenger mechanism in many systems, much remains unknown about their mechanism of action, except that such events are associated with increases or decreases in intracellular cyclic nucleotides. Quantitative descriptions of cyclic nucleotide-dependent processes are hampered by the absence of a means by which intracellular cyclic nucleotide concentrations can be accurately controlled. We have now designed, synthesized and characterized new, substituted photolabile cyclic nucleotide analogues, the 4,5-dimethoxy-2-nitrobenzyl esters of cyclic AMP and cyclic GMP (Fig. 1), which are physiologically inert before irradiation and which liberate free cyclic AMP or cyclic GMP on absorption of a photon. The thermal properties and photolysis rates and efficiencies of light-induced release of cyclic nucleotides from these analogues are more favourable than for the simple o-nitrobenzyl derivatives used previously. These molecules should permit intracellular "concentration jumps' of cyclic AMP or cyclic GMP to be produced in cells under physiological investigation with spatial and temporal resolution unmatched by conventional techniques.

Chemical Phenomena↗
PubMedDate not supplied

GABA uptake and release by a mammalian cell line stably expressing a cloned rat brain GABA transporter.

In order to facilitate study of the neuronal GABA transporter and provide a convenient system for potential drug screening, we have established a CHO cell line, designated 1F9, which stably expresses the cloned GABA transporter from rat brain (GAT-1). 1F9 cells transport GABA at levels approximately 300-fold higher than untransfected CHO cells, and GABA transport in these cells has the following properties: (1) a dependence on sodium and chloride ions; (2) higher sensitivity to neuronal subtype uptake inhibitors (DABA and ACHC) than to glial subtype inhibitors (beta-alanine and THPO); and (3) Km (2.5 microM) and IC50 values for various competitive ligands that are comparable with values determined in synaptosomes and brain slices. Given the fidelity with which the 1F9 cell line expresses these characteristics of the native neuronal GABA transporter, we have used it to further address GABA transporter activity. [3H]GABA uptake by 1F9 cells is inhibited approximately 50% by the chloride transport blockers DIDS and SITS. The GABA receptor agonists muscimol and baclofen also inhibit GABA transport; however, the receptor antagonists bicuculline and phaclofen have no effect. 1F9 cells also show release of [3H]GABA release is calcium independent, and is differentially affected by changes in the ion gradient, as well as by the presence of external substrates and uptake blockers. These experiments indicate that 1F9 cells provide a convenient system for the screening of GABA transport inhibitors.

Animals↗