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S Misler

Publications and source records attributed to S Misler.

At least 37 records · Page 2Linked to original sources
PubMed1992

Electrophysiology of stimulus-secretion coupling in human beta-cells.

Herein, we review the applicability to human beta-cells of an electrophysiologically based hypothesis of the coupling of glucose metabolism to insulin secretion. According to this hypothesis, glucose metabolism leads to the generation of intracellular intermediates (including ATP), which leads to closure of ATP-sensitive K+ channels. Channel closure results in membrane depolarization, the onset of electrical activity, and voltage-dependent Ca2+ entry. The resultant rise in cytosolic Ca2+ leads to Ca(2+)-dependent exocytosis of insulin granules. We found that most of the published experimental evidence for human beta-cells supports this hypothesis. In addition, we present three other emerging lines of evidence in support of this hypothesis for human islet beta-cells: 1) the effects of pHi-altering maneuvers on insulin secretion and electrical activity; 2) preliminary identification of LVA and HVA single Ca2+ channel currents; and 3) validation of the feasibility of Cm measurements to track insulin granule exocytosis. On the basis of this last new line of evidence, we suggest that combinations of Cm measurements and electrical activity/membrane current measurements may help define the roles of diverse electrical activity patterns, displayed by human beta-cells, in stimulus-induced insulin secretion.

Animals↗
PubMed1992

Stimulus-secretion coupling in beta-cells of transplantable human islets of Langerhans. Evidence for a critical role for Ca2+ entry.

With human islets isolated for transplantation, we examined the applicability to humans of a metabolic fuel hypothesis of glucose transduction and a Ca2+ hypothesis of depolarization-secretion coupling, both previously proposed for rodent islet beta-cells. We report that several features of human beta-cell physiology are well accounted for by these hypotheses. With whole-islet perifusion, we demonstrated that insulin secretion induced by glucose, tolbutamide, or elevated K+ is dependent on extracellular Ca2+. Insulin release induced by these secretagogues is enhanced by the dihydropyridine Ca2+ channel agonist BAYk8644 and depressed by the dihydropyridine Ca(2+)-channel antagonist nifedipine. All of the aforementioned secretagogues provoke increases in cytosolic free Ca2+, which are dependent on extracellular Ca2+ and are altered by the dihydropyridine drugs. Individual beta-cells in the islet display diminished resting membrane conductance, graded depolarization, and complex electrical patterns, including bursts of action potentials in response to stimulatory concentrations of glucose or tolbutamide. Individual islet beta-cells display voltage-dependent Ca2+ currents that are activated at membrane potentials traversed during the excursion of the action potential. In most cells, the Ca2+ currents are enhanced by BAYk8644 and depressed by nifedipine at concentrations that have parallel effects on secretagogue-induced increases in cytosolic Ca2+ and insulin secretion. These survey studies should provide the basis for more detailed investigations of the relationship of voltage-dependent ionic currents to electrical activity patterns and of electrical activity patterns to granule exocytosis in single human beta-cells.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1991

Single cell assay of exocytosis from adrenal chromaffin cells using "perforated patch recording".

We have combined "perforated patch recording" with phase detection to examine depolarization-induced changes in membrane capacitance (delta Cm) in bovine adrenal chromaffin cells. With this technique, voltage dependent Ca2+ currents and resultant delta Cm's often show little rundown over 1-2 hours even when the free Ca2+ concentration of the pipette is in the millimolar range. By limiting washout of cytosolic components and by maintaining more intact cytosolic Ca2+ buffering, this approach should facilitate the study of stimulus-exocytosis coupling evoked by physiological stimuli which involve cell metabolism and/or membrane receptor triggered second messenger cascades.

Adrenal Medulla↗
PubMed1991

Role of voltage-dependent ionic currents in coupling glucose stimulation to insulin secretion in canine pancreatic islet B-cells.

Glucose-induced electrical activity in canine pancreatic islet B cells is distinct from that in rodent islets, though both display Ca(2+)-dependent insulin secretion. Rodent islet B cells undergo regular bursts of Ca(2+)-dependent action potentials, while canine islet B cells generate isolated Na(+)-dependent action potentials which often give way to a plateau depolarization. Here we present evidence to reconcile the species difference in electrical activity with the similarity of Ca2+ dependence of secretion. (i) In canine B cells increasing glucose concentrations produce membrane depolarization and increasing frequency of Nao-dependent action potentials until a background membrane potential (approximately -40 mV) is reached where Na+ currents are inactivated. (ii) Voltage-dependent Ca2+ currents are present which are activated over the voltage excursion of the action potential (-50 to +20 mV) and inactivate slowly, (over seconds) in the range of the plateau depolarization (-40 to -25 mV). Hence, they are available to contribute to both phases of depolarization. (iii) Tetrodotoxin (TTX) reduces by half an early transient phase of glucose-stimulated insulin secretion but not a subsequent prolonged plateau phase. The transient phase of secretion often corresponds well in time to the period of initial high frequency action potential activity. These latter results suggest that in canine B cells voltage-dependent Na+ and Ca2+ currents mediate biphasic glucose-induced insulin secretion. The early train of Na(+)-dependent action potentials, by transiently activating Ca2+ channels and allowing pulsatile Ca2+ entry, may promote an early transient phase of insulin secretion. The subsequent sustained plateau depolarization, by allowing sustained Ca2+ entry, may permit steady insulin release.

Action Potentials↗
PubMed1991

Long-term monitoring of depolarization-induced exocytosis from adrenal medullary chromaffin cells and pancreatic islet B cells using "perforated patch recording".

(1) Membrane capacitance measurements using perforated patch recording offer the possibility of studying the process of depolarization-secretion coupling (DSC) in single endocrine cells with unprecedented time resolution and stability. (2) Early results with catecholamine-secreting adrenal chromaffin cells and insulin-secreting pancreatic B cells support longstanding ideas that the Ca(2+)-dependent processes underlying DSC are fundamentally similar to those of nerve terminals. (3) Future experiments using these approaches should prove useful in sorting out those effects of humoral substances that have a predominant effect on excitability and Ca2+ entry from those that affect the secretory process itself.

Adrenal Medulla↗
PubMed1990

Sodium channels contribute to action potential generation in canine and human pancreatic islet B cells.

Pancreatic islet B cells depolarize and display trains of action potentials in response to stimulatory concentrations of glucose. Based on data from rodent islets these action potentials are considered to be predominantly Ca2+ dependent. Here we describe Na(+)-dependent action potentials and Na+ currents recorded from canine and human pancreatic islet B cells. Current-clamp recording using the nystatin "perforated-patch" technique demonstrates that B cells from both species display tetrodotoxin-sensitive Na+ action potentials in response to modest glucose-induced depolarization. In companion "whole-cell" voltage-clamp experiments on canine B cells, the underlying Na+ current displays steep voltage-dependent activation and inactivation over the range of -50 to -40 mV. The Na+ current is sensitive to tetrodotoxin block with a KI = 3.2 nM and has a reversal potential which changes with [Na+]o as predicted by the Nernst equation. These results suggest that a voltage-dependent Na+ current may contribute significantly to action potential generation in some species outside the rodent family.

Action Potentials↗
PubMed1989

Voltage-activated and stretch-activated Ba2+ conducting channels in an osteoblast-like cell line (UMR 106).

Calcium entry may mediate osteoblast activation by calciotropic hormones or changes in electrical fields or mechanical stress in bone. Using cell-attached patch clamping techniques, we have seen two Ba2+ conducting channels in UMR-106 osteoblast-like cells: (i) a voltage-dependent, dihydropyridine-sensitive channel resembling L-type Ca2+ channels in other cells, and (ii) a stretch-activated non-selective channel resembling those involved in mechanoreception and triggering of volume regulation in other cells.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1989

'Perforated patch recording' allows long-term monitoring of metabolite-induced electrical activity and voltage-dependent Ca2+ currents in pancreatic islet B cells.

We describe the application of 'perforated patch recording' using the pore-forming antibiotic nystatin, to monitor the electrical activity and underlying ionic currents of rat and human pancreatic islet B cells. We demonstrate that glucose-induced electrical activity is seen even in single B cells during current-clamp recordings lasting hours 'L-type' Ca2+-channel currents can also be monitored over this period of time. This technique may prove useful in examining hormone and neurotransmitter modulation of electrical activity in B cells, while minimizing the effects of cytoplasmic 'wash-out'.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1989

Ca2+-activated K+ channel in rat pancreatic islet B cells: permeation, gating and blockade by cations.

Activation of Ca2+-dependent K+ conductance has long been postulated to contribute to the cyclical pauses in glucose-induced electrical activity of pancreatic islet B cells. Here we have examined the gating, permeation and blockade by cations of a large-conductance, Ca2+-activated K+ channel in these cells. This channel shares many features with BK (or maxi-K+) Ca2+-activated K+ channels in other cells. (1) Its 'permeability' selectivity sequence is PT1+: PK+: PRb+: PNH4+: PNa+, Li+, Cs+ = 1.3:1.0:0.5:0.17: less than 0.05. Permeant, as well as impermeant, cations reduce channel conductance. (2) Its conductance saturates at 325-350 pS with bath KCl greater than 400 mM (144 mM KCl pipette). (3) It shows asymmetric blockade by tetraethylammonium ion (TEA) and Na+. (4) It is sensitive to Ca2+i over the range 5 nM-100 microM; over the range 50-200 nM, channel activity varies as [Ca2+ free]1-2. (5) It is sensitive to internal pH over the range 6.85-7.35, but the decrease in channel activity seen with reduced pHi may be partially compensated by the increase in free Ca2+ concentration which occurs on acidification of buffered Ca2+/EGTA solutions.

Animals↗
PubMed1989

Modulation of gating of a metabolically regulated, ATP-dependent K+ channel by intracellular pH in B cells of the pancreatic islet.

Membrane-permeant weak acids and bases, when applied to the bath, modulate the resting membrane potential and the glucose-induced electrical activity of pancreatic B cells, as well as their insulin secretion. These substances alter the activity of a metabolite-regulated, ATP-sensitive K+ channel which underlies the B-cell resting potential. We now present several lines of evidence indicating that the channel may be directly gated by pHi. (1) The time course of K+(ATP) channel activity during exposure to and washout of NH4Cl under a variety of experimental conditions, including alteration of the electrochemical gradient for NH4Cl entry and inhibition of the Na+o/H+i exchanger, resembles the time course of pHi measured in other cell types that have been similarly treated. (2) Increasing pHo over the range 6.25-7.9 increases K+(ATP) channel activity in cell-attached patches where the cell surface exposed to the bath has been permeabilized to H+ by the application of the K+/H+ exchanger nigericin. (3) Increasing pHi over a similar range produces similar effects on K+(ATP) channels in inside-out excised patches exposed to small concentrations of ATPi. The physiological role of delta pHi in the metabolic gating of this channel remains to be explored.

Adenosine Triphosphate↗
PubMed1989

Activity of ion channels during volume regulation by clonal N1E115 neuroblastoma cells.

When exposed to a hypotonic bathing solution, clonal N1E115 neuroblastoma cells initially swell and then undergo a regulatory volume decrease (RVD). Using cell-attached patch-clamp recording, we have found that the activity of a stretch-sensitive, nonselective cation [C+(SA)] channel increases shortly after the onset of osmotically induced cell swelling; this depolarizes the cells as much as 30 mV. Shortly thereafter, and roughly coincident with the onset of RVD, two types of voltage-dependent channels open at the new resting potential; these are (i) a delayed-rectifier type K+ [K+(DR)] channel and (ii) a large-conductance anion channel. We suggest that opening of the C+(SA) channel may contribute to the volume "sensor" mechanism, while the depolarization-induced opening of the K+(DR) and anion channels may constitute a significant K+ salt exit pathway, operating in RVD.

Animals↗
PubMed1989

Effects of sulfonamides on a metabolite-regulated ATPi-sensitive K+ channel in rat pancreatic B-cells.

Intracellular ATP (ATPi)-sensitive K+ [K+(ATP)] channels are now a recognized site of action of clinically useful hypoglycemic and hyperglycemic sulfonamides. We have further examined the action of these agents on single K+ channels in rat pancreatic B-cells 1) Tolbutamide and glyburide, two hypoglycemic sulfonylureas which decrease K+(ATP) channel activity in the cell-attached patch, affect the kinetics of K+(ATP) channel in a manner similar to glucose. They shorten the duration of the "burst," or cluster of open channel events, while lengthening the intervals between bursts. 2) The hyperglycemic vasodilator diazoxide increases mean K+(ATP) channel activity in the cell-attached patch as well as in the inside-out excised patch exposed to ATPi. It appears to lengthen channel bursts and shorten the intervals between them. Two structurally similar diuretics, hydrochlorothiazide and furosemide, which have mild hyperglycemic effects, do not increase K+(ATP) channel activity even at clinically toxic concentrations. 3) Neither the sulfonylureas nor diazoxide directly affect the activity of single delayed rectifier K+ channels or single calcium and voltage-activated K+ channels in normal B-cells.

Adenosine Triphosphate↗
PubMed1989

Metabolite-regulated ATP-sensitive K+ channel in human pancreatic islet cells.

In patch-clamped surface cells of human islets, we identified an inwardly rectifying, voltage-independent K+ channel that may be a crucial link between substrate metabolism and depolarization-induced insulin secretion. It is the major channel open at rest. It closes on exposure of the cell to secretagogue concentrations of glucose or other metabolic fuels and oral hypoglycemic sulfonylureas but reopens on addition of either a metabolic inhibitor that prevents substrate utilization or the hyperglycemic sulfonamide diazoxide. Onset of electrical activity coincides with channel closure by the secretagogues. In excised patches, the activity of this channel is inhibited at its cytoplasmic surface by ATP. These results suggest that in humans, as in rodents, 1) rises in cytoplasmic ATP levels during substrate metabolism trigger K+-channel closure and cell depolarization and 2) clinically useful sulfonamides modulate glucose-induced insulin secretion, in part by affecting a readily identifiable resting conductance pathway for K+.

Action Potentials↗
PubMed1988

A stretch-activated anion channel in tobacco protoplasts.

Stretch-activated ion channels have been described in animal cells, where they might serve as mechanoreceptors, baroreceptors or osmoreceptors, as well as in yeast and bacteria, where osmoregulatory functions have been suggested. Here we report a large conductance, stretch-activated, anion-selective channel in protoplasts of a higher plant, tobacco, and discuss its possible role in osmoregulation.

Ion Channels↗
PubMed1987

Cation dependence of posttetanic potentiation of neuromuscular transmission.

We have investigated the possibility that much of posttetanic potentiation (PTP) of quantal release of neurotransmitter at the frog neuromuscular junction may be due to posttetanic accumulation of [Ca2+]i, via a plasmalemmal Cao2+-Nai+ exchanger that is powered by an increase in Nai+ during the tetanus. Our new findings on the cationic dependence of PTP are consistent with this hypothesis. 1) Several manuevers that decrease Na+-K+ pump activity, (decreasing [K+]o, replacing K+o with Rb+o or Li+o, or adding acetylstrophanthidin to Ringers), all increase the intratetanic rise and prolong the posttetanic decay of epp quantal content (m) and miniature epp frequency (fmepp). 2) Increasing [Ca2+]i or [Sr2+]o, but not [Mg2+]o, increases posttetanic fmepp in a graded fashion. 3) PTP of fmepp is still present after addition of Mn2+o, which blocks voltage dependent Ca2+ entry.

Animals↗
PubMed1987

Dependence on multivalent cations of quantal release of transmitter induced by black widow spider venom.

Application of alpha-latrotoxin (alpha-LT), the active component of black widow spider venom (BWSV), to a vertebrate neuromuscular junction, in the presence of millimolar bath concentrations of Ca2+ or Mg2+, greatly increases the frequency of miniature end-plate potentials (Fmepp). We have further characterized the cation dependence of alpha-LT action at the frog cutaneous pectoris neuromuscular junction. The divalent cations, Ca, Sr, Ba at less than or equal to 50 microM, Zn, Mn, Cd at greater than or equal to 50-100 microM, and Mg at greater than or equal to 1.0 mM, as well as the trivalent cation La at greater than or equal to 15 microM, all increase Fmepp exponentially to greater than or equal to 100-200 s-1 over several minutes time. The exponential rate of rise is graded with extracellular cation concentration and can be reduced by increasing [K+] of the bath from 2 to 25-40 mM. Long-term exposure to alpha-LT in the presence of Sr2+ or Mn2+ results in the exhaustion of the releasable quantal store of transmitter, which in the case of Mn2+ correlates well with depletion of synaptic vesicles. These data support the hypothesis that BWSV promotes an increase in Fmepp by increasing nerve terminal permeability to multivalent cations that enter the nerve terminal down their electrochemical gradients and then may bind to quantal release activating sites or displace Ca2+ from intracellular stores.

Animals↗
PubMed1986

A metabolite-regulated potassium channel in rat pancreatic B cells.

In B cells from dispersed rat islet of Langerhans we have identified an inward rectifying voltage-independent K+ channel whose behavior parallels the metabolically regulated potassium permeability (PK) found in tracer flux and microelectrode recording studies. In cell-attached patches of membrane, the channel is closed when any one of several substrates (glucose, mannose, leucine, or glyceraldehyde) is added to the cell's bathing solution but is reopened on addition of an appropriate metabolic inhibitor, which prevents utilization of that substrate. In inside-out excised patches, a K+ channel with nearly identical kinetic features is closed by addition of micromolar concentrations of ATP to the "cytoplasmic" solution. The ATP sensitivity of channel activity is modified by addition of ADP, suggesting competition at a nucleotide binding site. These results suggest the presence of a metabolically regulated K+ channel gated by intracellular concentrations of ATP or the ratio of ATP/ADP concentrations.

Adenosine Diphosphate↗