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Cell calcium and the control of membrane transport : Society of General Physiologists, 40th Annual Symposium : Marine Biological Laboratory, Woods Hole, Massachusetts, 3-7 September, 1986
01 Jan 1987-
TL;DR: A compendium of the written contributions submitted by the invited speakers to the 40th Annual Meeting of the Society of General Physiologists can be found in this paper, where the authors also published 118 abstracts of contributed papers submitted to this meeting.
Abstract: Abstract : This book is a compendium of the written contributions submitted by the invited speakers to the 40th Annual Meeting of the Society of General Physiologists. There were also 118 abstracts of contributed papers submitted to this meeting, which have been published in the December 1986 issue of The Journal of General Physiology. Partial Contents: Regulation of Cytosolic Free Calcium, The Plasma Membrane in the Control of the Signaling Function of Calcium, Calcium-permeable Channels in Vascular Smooth Muscle: Voltage-activated, Receptor-operated, and Leak Channels, Calcium and Magnesium Movements in Cells and the Role of Inositol Trisphosphate in Muscle, Receptor-mediated Changes in Intracellular Calcium, Mechanisms Involved in Receptor-mediated Changes of Intracellular Ca2 + in Liver, The Role of Phosphatidylinositides in Stimulus-Secretion Coupling in the Exocrine Pancreas, Modulation of Membrane Transport by Intracellular Calcium, The Role of Cyclic AMP-dependent Phosphorylation in the Maintenance and Modulation of Voltage-activated Calcium Channels, Multiple Roles for Calcium and Calcium-dependent Enzymes in the Activation of Peptidergic Neurons of Aplysia, Calcium Involvement in Intracellular Events, The Relationship Between the Cytosolic Free Calcium Ion Concentration and the Control of Pyruvate Dehydrogenase, Membrane and Microfilament Organization and Vasopressin Action in Transporting Epithelia.
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TL;DR: The proposition that depolarized hippocampal mossy fiber synaptosomes release endogenous ATP and are capable of forming adenosine from extracellular ATP is supported, and that endogenousAdenosine may act at a presynaptic site to inhibit the further release of glutamic acid and the prodynorphin‐derived peptides is supported.
Abstract: Using a hippocampal subcellular fraction enriched in mossy fiber synaptosomes, evidence was obtained indicating that adenosine derived from a presynaptic pool of ATP may modulate the release of prodynorphin-derived peptides and glutamic acid from mossy fiber terminals. Synaptosomal ATP was released in a Ca2+-dependent manner by K+-in-duced depolarization. The rapid hydrolysis of extracellular [14C]ATP in the presence of intact mossy fiber synaptosomes resulted in the production of [14C]adenosine. Micromolar concentrations of a stable adenosine analogue, 2-chloro-adenosine, inhibited the K+-stimulated release of both dynorphin B and dynorphin A(l-8). 2-Chloroadenosine failed to suppress the evoked release of glutamic acid, measured in these same superfusates, unless the mossy fiber synaptosomes were pretreated with D-aspartic acid to deplete the cytosolic, Ca2+-independent, pool of this acidic amino acid. In synaptosomes pretreated in this manner, release of the remaining Ca2+-dependent pool of glutamic acid was significantly inhibited by NiCl2, 2-chloroadenosine, 5′-N-ethylcarboxami-doadenosine, cyclohexyladenosine, and R(-)-N6(2-phenyl-isopropyl)adenosine, but not by ATP. 2-Chloroadenosine-induced inhibition was reversed when the external CaCl2concentration was raised from 1.8 mMto 6 mM. 8-Phen-yltheophylline, an adenosine receptor antagonist, effectively blocked the inhibitory effects of 2-chloroadenosine on mossy fiber synaptosomes and significantly enhanced the K+-evoked release of both glutamic acid and dynorphin A(l-8) when added alone to the superfusion medium. These results support the proposition that depolarized hippocampal mossy fiber synaptosomes release endogenous ATP and are capable of forming adenosine from extracellular ATP, and that endogenous adenosine may act at a presynaptic site to inhibit the further release of glutamic acid and the prodynorphin-derived peptides
91 citations
TL;DR: The inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells only occurs when membrane depolarization leads to calcium ion entry and intracellular accumulation.
Abstract: The inactivation of calcium channels in mammalian pituitary tumor cells (GH3) was studied with patch electrodes under voltage clamp in cell-free membrane patches and in dialyzed cells The calcium current elicited by depolarization from a holding potential of -40 mV passed predominantly through one class of channels previously shown to be modulated by dihydropyridines and cAMP-dependent phosphorylation (Armstrong and Eckert, 1987) When exogenous calcium buffers were omitted from the pipette solution, the macroscopic calcium current through those channels inactivated with a half time of approximately 10 ms to a steady state level 40-75% smaller than the peak Inactivation was also measured as the reduction in peak current during a test pulse that closely followed a prepulse Inactivation was largely reduced or eliminated by (a) buffering free calcium in the pipette solution to less than 10(-8) M; (b) replacing extracellular calcium with barium; (c) increasing the prepulse voltage from +10 to +60 mV; or (d) increasing the intracellular concentration of cAMP, either 'directly' with dibutyryl-cAMP or indirectly by activating adenylate cyclase with forskolin or vasoactive intestinal peptide Thus, inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells only occurs when membrane depolarization leads to calcium ion entry and intracellular accumulation
77 citations
TL;DR: The data suggest that steady state [Ca2+]i in CCT cells resulted from a non-saturable passive entry of calcium ions across cell membranes balanced with an active extrusion by calcium ATPase (pump and leak mechanism).
Abstract: Cytosolic free Ca2+ ([Ca2+]i) was measured in single fragments of rat cortical collecting tubule (CCT) by using fura-2 and a tubule superfusion device. Under basal conditions, i.e. with 1 mM of external Ca2+ ([Ca2+]o), the average steady state [Ca2+]i was 179±16 nM (n=44 tubules). Random alterations of [Ca2+]o between 0 mM and 4 mM led to corresponding variations in steady state [Ca2+]i levels, which were linearly correlated with [Ca2+]o (average slope 93±34 nM [Ca2+]i per 1 mM [Ca2+]o for six tubules). In contrast, [Ca2+]i was little affected by decreasing external Na+ concentration. Cell membrane depolarization with 100 mM of external K+ induced a sustained drop in [Ca2+]i (21% as an average). The data suggest that steady state [Ca2+]i in CCT cells resulted from a non-saturable passive entry of calcium ions across cell membranes balanced with an active extrusion by calcium ATPase (pump and leak mechanism). The passive component cannot be accounted for either by Na+/Ca2+ exchangers nor by voltage-dependent calcium channels; it is best explained by the presence of voltage-independent calcium channels in cell membranes.
61 citations
TL;DR: It is suggested that Ihyp is mediated by a novel, hyperpolarization-activated calcium conductance that is distinct from the one activated by depolarization.
Abstract: Hyperpolarization of Paramecium tetraurelia under conditions where K+ currents are suppressed elicits an inward current that activates rapidly toward a peak at 25-80 ms and decays thereafter. This peak current (Ihyp) is not affected by removing Cl ions from the microelectrodes used to clamp membrane potential, or by changing extracellular Cl- concentration, but is lost upon removing extracellular Ca2+. Ihyp is also lost upon replacing extracellular Ca2+ with equimolar concentrations of Ba2+, Co2+, Mg2+, Mn2+, or Sr2+, suggesting that the permeability mechanism that mediates Ihyp is highly selective for Ca2+. Divalent cations also inhibit Ihyp when introduced extracellularly, in a concentration- and voltage-dependent manner. Ba2+ inhibits Ihyp with an apparent dissociation constant of 81 microM at -110 mV, and with an effective valence of 0.42. Ihyp is also inhibited reversibly by amiloride, with a dissociation constant of 0.4 mM. Ihyp is not affected significantly by changes in extracellular Na+, K+, or H+ concentration, or by EGTA injection. Also, it is unaffected by manipulations or mutations that suppress the depolarization-activated Ca2+ current or the various Ca(2+)-dependent currents of Paramecium. We suggest that Ihyp is mediated by a novel, hyperpolarization-activated calcium conductance that is distinct from the one activated by depolarization.
60 citations
TL;DR: The results demonstrate thatCa2+ flux into dentinogenically active odontoblasts occurs through voltage-gated Ca2+ channels of the L-type and by nonvoltage-gating, agonist-sensitive Ca2-sensitive uptake pathways.
Abstract: Odontoblasts participate actively in the transport and accumulation of Ca2+ ions to the mineralization front during dentinogenesis. These cells are known to carry membrane-bound ATP-driven pumps and Na+/Ca2+ antiports for Ca2+ extrusion, but little is known about Ca2+ influx mechanisms into these cells. It has been shown that the administration of Ca2+ channel blockers in vivo strongly impairs Ca2+ uptake in the mineral phase during dentinogenesis in the rat; the present in vitro study is aimed at further elucidating odontoblast Ca2+ uptake mechanisms. Dissected rat incisor odontoblasts exhibited a pronounced fluorescence when incubated with a fluorescently-labeled (STBodipy) dihydropyridine, which is specific for voltage-gated Ca2+ channels of the L-type, and this binding was competitively abolished by nifedipine. As assayed by fluorescence spectrometry, odontoblast Ca2+ uptake was enhanced by the agonistic dihydropyridine BAYK-8644 (5 μM) as well as by plasma membrane depolarization in a high K+ (120 mM) medium. The Ca2+ uptake after depolarization was impaired by nifedipine (5 μM). When treated with the Ca2+-ATPase inhibitor cyclopiazonic acid (CPA; 10 μM), a nonvoltagegated uptake of 45Ca2+ was identified. This uptake was not influenced by nifedipine (20 μM) but was impaired by lanthanum ions (200 μM). A nonvoltage-gated uptake of Mn2+ into CPA-treated cells could be traced using the fura-2 quenching technique. This CPA-induced Ca2+ flux was not caused by an alteration of the plasma membrane potential, as assayed with di-8-ANEPPS. The results demonstrate that Ca2+ flux into dentinogenically active odontoblasts occurs through voltage-gated Ca2+ channels of the L-type and by nonvoltage-gated, agonist-sensitive Ca2+ uptake pathways.
40 citations
References
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TL;DR: The proposition that depolarized hippocampal mossy fiber synaptosomes release endogenous ATP and are capable of forming adenosine from extracellular ATP is supported, and that endogenousAdenosine may act at a presynaptic site to inhibit the further release of glutamic acid and the prodynorphin‐derived peptides is supported.
Abstract: Using a hippocampal subcellular fraction enriched in mossy fiber synaptosomes, evidence was obtained indicating that adenosine derived from a presynaptic pool of ATP may modulate the release of prodynorphin-derived peptides and glutamic acid from mossy fiber terminals. Synaptosomal ATP was released in a Ca2+-dependent manner by K+-in-duced depolarization. The rapid hydrolysis of extracellular [14C]ATP in the presence of intact mossy fiber synaptosomes resulted in the production of [14C]adenosine. Micromolar concentrations of a stable adenosine analogue, 2-chloro-adenosine, inhibited the K+-stimulated release of both dynorphin B and dynorphin A(l-8). 2-Chloroadenosine failed to suppress the evoked release of glutamic acid, measured in these same superfusates, unless the mossy fiber synaptosomes were pretreated with D-aspartic acid to deplete the cytosolic, Ca2+-independent, pool of this acidic amino acid. In synaptosomes pretreated in this manner, release of the remaining Ca2+-dependent pool of glutamic acid was significantly inhibited by NiCl2, 2-chloroadenosine, 5′-N-ethylcarboxami-doadenosine, cyclohexyladenosine, and R(-)-N6(2-phenyl-isopropyl)adenosine, but not by ATP. 2-Chloroadenosine-induced inhibition was reversed when the external CaCl2concentration was raised from 1.8 mMto 6 mM. 8-Phen-yltheophylline, an adenosine receptor antagonist, effectively blocked the inhibitory effects of 2-chloroadenosine on mossy fiber synaptosomes and significantly enhanced the K+-evoked release of both glutamic acid and dynorphin A(l-8) when added alone to the superfusion medium. These results support the proposition that depolarized hippocampal mossy fiber synaptosomes release endogenous ATP and are capable of forming adenosine from extracellular ATP, and that endogenous adenosine may act at a presynaptic site to inhibit the further release of glutamic acid and the prodynorphin-derived peptides
91 citations
TL;DR: The inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells only occurs when membrane depolarization leads to calcium ion entry and intracellular accumulation.
Abstract: The inactivation of calcium channels in mammalian pituitary tumor cells (GH3) was studied with patch electrodes under voltage clamp in cell-free membrane patches and in dialyzed cells The calcium current elicited by depolarization from a holding potential of -40 mV passed predominantly through one class of channels previously shown to be modulated by dihydropyridines and cAMP-dependent phosphorylation (Armstrong and Eckert, 1987) When exogenous calcium buffers were omitted from the pipette solution, the macroscopic calcium current through those channels inactivated with a half time of approximately 10 ms to a steady state level 40-75% smaller than the peak Inactivation was also measured as the reduction in peak current during a test pulse that closely followed a prepulse Inactivation was largely reduced or eliminated by (a) buffering free calcium in the pipette solution to less than 10(-8) M; (b) replacing extracellular calcium with barium; (c) increasing the prepulse voltage from +10 to +60 mV; or (d) increasing the intracellular concentration of cAMP, either 'directly' with dibutyryl-cAMP or indirectly by activating adenylate cyclase with forskolin or vasoactive intestinal peptide Thus, inactivation of the dihydropyridine-sensitive calcium channels in GH3 cells only occurs when membrane depolarization leads to calcium ion entry and intracellular accumulation
77 citations
TL;DR: The data suggest that steady state [Ca2+]i in CCT cells resulted from a non-saturable passive entry of calcium ions across cell membranes balanced with an active extrusion by calcium ATPase (pump and leak mechanism).
Abstract: Cytosolic free Ca2+ ([Ca2+]i) was measured in single fragments of rat cortical collecting tubule (CCT) by using fura-2 and a tubule superfusion device. Under basal conditions, i.e. with 1 mM of external Ca2+ ([Ca2+]o), the average steady state [Ca2+]i was 179±16 nM (n=44 tubules). Random alterations of [Ca2+]o between 0 mM and 4 mM led to corresponding variations in steady state [Ca2+]i levels, which were linearly correlated with [Ca2+]o (average slope 93±34 nM [Ca2+]i per 1 mM [Ca2+]o for six tubules). In contrast, [Ca2+]i was little affected by decreasing external Na+ concentration. Cell membrane depolarization with 100 mM of external K+ induced a sustained drop in [Ca2+]i (21% as an average). The data suggest that steady state [Ca2+]i in CCT cells resulted from a non-saturable passive entry of calcium ions across cell membranes balanced with an active extrusion by calcium ATPase (pump and leak mechanism). The passive component cannot be accounted for either by Na+/Ca2+ exchangers nor by voltage-dependent calcium channels; it is best explained by the presence of voltage-independent calcium channels in cell membranes.
61 citations
TL;DR: It is suggested that Ihyp is mediated by a novel, hyperpolarization-activated calcium conductance that is distinct from the one activated by depolarization.
Abstract: Hyperpolarization of Paramecium tetraurelia under conditions where K+ currents are suppressed elicits an inward current that activates rapidly toward a peak at 25-80 ms and decays thereafter. This peak current (Ihyp) is not affected by removing Cl ions from the microelectrodes used to clamp membrane potential, or by changing extracellular Cl- concentration, but is lost upon removing extracellular Ca2+. Ihyp is also lost upon replacing extracellular Ca2+ with equimolar concentrations of Ba2+, Co2+, Mg2+, Mn2+, or Sr2+, suggesting that the permeability mechanism that mediates Ihyp is highly selective for Ca2+. Divalent cations also inhibit Ihyp when introduced extracellularly, in a concentration- and voltage-dependent manner. Ba2+ inhibits Ihyp with an apparent dissociation constant of 81 microM at -110 mV, and with an effective valence of 0.42. Ihyp is also inhibited reversibly by amiloride, with a dissociation constant of 0.4 mM. Ihyp is not affected significantly by changes in extracellular Na+, K+, or H+ concentration, or by EGTA injection. Also, it is unaffected by manipulations or mutations that suppress the depolarization-activated Ca2+ current or the various Ca(2+)-dependent currents of Paramecium. We suggest that Ihyp is mediated by a novel, hyperpolarization-activated calcium conductance that is distinct from the one activated by depolarization.
60 citations
TL;DR: The results demonstrate thatCa2+ flux into dentinogenically active odontoblasts occurs through voltage-gated Ca2+ channels of the L-type and by nonvoltage-gating, agonist-sensitive Ca2-sensitive uptake pathways.
Abstract: Odontoblasts participate actively in the transport and accumulation of Ca2+ ions to the mineralization front during dentinogenesis. These cells are known to carry membrane-bound ATP-driven pumps and Na+/Ca2+ antiports for Ca2+ extrusion, but little is known about Ca2+ influx mechanisms into these cells. It has been shown that the administration of Ca2+ channel blockers in vivo strongly impairs Ca2+ uptake in the mineral phase during dentinogenesis in the rat; the present in vitro study is aimed at further elucidating odontoblast Ca2+ uptake mechanisms. Dissected rat incisor odontoblasts exhibited a pronounced fluorescence when incubated with a fluorescently-labeled (STBodipy) dihydropyridine, which is specific for voltage-gated Ca2+ channels of the L-type, and this binding was competitively abolished by nifedipine. As assayed by fluorescence spectrometry, odontoblast Ca2+ uptake was enhanced by the agonistic dihydropyridine BAYK-8644 (5 μM) as well as by plasma membrane depolarization in a high K+ (120 mM) medium. The Ca2+ uptake after depolarization was impaired by nifedipine (5 μM). When treated with the Ca2+-ATPase inhibitor cyclopiazonic acid (CPA; 10 μM), a nonvoltagegated uptake of 45Ca2+ was identified. This uptake was not influenced by nifedipine (20 μM) but was impaired by lanthanum ions (200 μM). A nonvoltage-gated uptake of Mn2+ into CPA-treated cells could be traced using the fura-2 quenching technique. This CPA-induced Ca2+ flux was not caused by an alteration of the plasma membrane potential, as assayed with di-8-ANEPPS. The results demonstrate that Ca2+ flux into dentinogenically active odontoblasts occurs through voltage-gated Ca2+ channels of the L-type and by nonvoltage-gated, agonist-sensitive Ca2+ uptake pathways.
40 citations