Transmembrane Calcium Flux Research Articles

Unwinding the Snail's Clock: Cellular Analysis of a Retinal Circadian Pacemaker

The eye of the cloudy bubble snail, Bulla gouldiana, expresses a circadian rhythm in spontaneous optic nerve impulse frequency. The circadian rhythm is generated among a group of approximately 100 neurons (basal retinal neurons - BRNs) at the base of the retina. Each BRN appears to be a competent circadian pacemaker. Using the Bulla retina as a model pacemaker system our laboratory has addressed three central issues of circadian physiology: Synchronization: Entrainment of the ocular rhythm to light cycles is mediated by depolarization of pacemaker neurons with a resultant transmembrane calcium flux. Phase shifts of the rhythm are prevented when calcium entry or membrane depolarization is blocked and light-induced phase shifts can be mimicked by depolarization. Depolarization leads to a persistent increase in intraccllular calcium in pacemaker neurons. Expression: The circadian rhythm in impulse frequency is driven by a rhythm in membrane conductance. Membrane conductance is relatively high during the subjective night and decreases by approximately 50 percent near subjective dawn. The 'clock-controlled' conductances are TEA sensitive and recent experiments reveal a circadian modulation in a sustained calcium-inclependent potassium conductance. Rhythm generation: Protein synthesis and transcription appear to play critical roles in rhythm genesis. Alterations in the rates of transcription and translation profoundly affect the velocity of the circadian pacemaker in a phase-dependent manner. Transmembrane ionic fluxes, while not involved in rhythm generation, play critical roles in pacemaker synchronization and rhythm expression.

Evidence that the antiproliferative effect of verapamil on afferent and efferent immune responses is independent of calcium channel inhibition.

Calcium channel blockers are capable of inhibiting the afferent and efferent limbs of the immune responses of human peripheral blood mononuclear cells in in vitro systems. This effect is thought to be related to the ability of the calcium channel blocker to limit the transmembrane flux of calcium. We report herein that two optical enantiomers of verapamil, one (S-) which is capable of blocking the slow calcium channel and mitogen-stimulated 45Ca++ uptake into human lymphocytes, while the other (R+) is incapable of either activity, share almost identical capabilities of depressing both the afferent and efferent limbs of immunity. These observations suggest that the inhibitory effects of verapamil on various afferent and efferent immune events are, in part at least, unrelated to the inhibition of transmembrane calcium flux.

Sialic acid removal modules the myocardial and vascular activity of calcium channel ligands

Selective removal of sialic acid from isolated guinea pig left atrial strips and rabbit thoracic aortic ring segments was performed by neuraminidase prepared from Clostridium perfringens and was controlled electron microscoplcally. Preincubation of these organs (2 units/mL; 2 hr) resulted in enzyme mediated hydrolysis of total tissue sialic acid; 55.2% for atria and 60.9% for aorta. Contractile force of atria and arterial diameter of thoracic aorta were measured isometrically and isotonically by means of a force displacement transducer. Pretreatment of both organs with neuraminidase (2 units/mL; 2 hr) in a carbogen saturated organ bath caused a moderate left-hand shift of the cumulative concentration response curves for the dihydropyridine type calcium antagonist nisoldipine, the phenylalkylamine derivative gallopamil and the benzothiazepine diltiazem. ec 50 values were significantly lower (P < 0.05), particularly in the atrial muscle, when compared to untreated preparations. There was no effect of neuraminidase on the negative inotroplc and vasodilator potency of the calcium channel modulator fendiline. Conversely, neuraminidase induced a right-hand shift in the concentration response curves shown by the pure calcium agonist (−)- S-Bay K 8644 leading to significantly higher ec 50 values in both organs. Similarly, the contractile potency of calcium chloride (atria) and potassium chloride (aorta) was attenuated upon neuraminidase treatment. From the results obtained it is concluded that sialic acid removal may modulate the action of calcium channel ligands through an inhibitory effect on transmembrane calcium fluxes and/or by decreasing the external calcium availability. Whether the present results suggest a functional role for sialic acid in the regulation of calcium channels warrants further investigation.

Induction of T cell CD7 gene transcription by nonmitogenic ionomycin-induced transmembrane calcium flux.

The CD7 molecule is a 40-kDa member of the Ig superfamily that has structural homology to the murine Thy-1 molecule and is acquired early in human T cell ontogeny. Previous studies have demonstrated that expression of the CD7 molecule is markedly up-regulated during T cell activation. In this study, we have studied the signals required for CD7 up-regulation on human T cells. We found that nonmitogenic amounts of ionomycin selectively and maximally up-regulated T cell CD7 on mature (peripheral blood) T cells after 24 h. Whereas CD7 expression was increased 78 +/- 25% by 0.5 microM ionomycin, expression of CD25 (IL-2R alpha), class II MHC, 4F2, transferrin receptor, CD2, CD3, CD4, CD5, and CD8 molecules was not increased. Ionomycin-induced CD7 surface expression was associated with peak increases in CD7 mRNA after 4 to 6 h. Transcriptional analysis and CD7 mRNA half-life determination revealed the increase in CD7 mRNA was the result of increased CD7 gene transcription 1 h after ionomycin stimulation and was not due to prolongation of CD7 mRNA half-life. The up-regulation of surface CD7 expression by ionomycin was dependent on extracellular calcium and did not require the activation of T cell tyrosine protein kinase. Mitogenic CD2 and CD3 mAb as well as stimulation of T cells by PHA also up-regulated CD7 expression. CD7 up-regulation by ionomycin was transient (24 to 72 h) and inhibitable by cyclosporin A, whereas CD7 up-regulation by PHA was sustained over 5 to 7 days and was significantly less inhibitable by cyclosporin A. These data demonstrate that induction of a transmembrane calcium flux generates signals that lead to CD7 gene transcription.

Cholecystokinin-induced phosphatidylinositol hydrolysis in rat pancreatic acinar cells: modulation by extracellular calcium and manganese.

The role of extracellular calcium in modulating the actions of cholecystokinin octapeptide (CCK8) on phosphatidylinositol 4,5-bisphosphate hydrolysis was studied in freshly isolated rat pancreatic acini and cultured AR42J cells. In both cell types, CCK8 rapidly induced the formation of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and 1,3,4, 5-tetrakisphosphate [Ins(1,3,4,5)P4]. The actions of CCK8 were inhibited by lanthanum and manganese, agents that block transmembrane calcium fluxes, and by chelation of extracellular calcium with EGTA. In pancreatic acini, lanthanum and manganese also partially inhibited the effects of carbachol and bombesin on Ins(1,4,5)P3 and Ins(1,3,4,5)P4 levels. In acini, the CCK8-mediated increases in Ins(1,4,5)P3 and Ins(1,3,4,5)P4 levels were progressively greater as the extracellular calcium concentration was raised from the micromolar range to 1.28 mM and progressively smaller as the manganese concentration was raised from 10 microM to 1 mM. Furthermore, the CCK8-mediated rise in Ins(1,4,5)P3 levels was partially attenuated by the calcium channel blockers verapamil, diltiazem, and nifedipine. These findings indicate that extracellular calcium enhances the ability of CCK8 and other calcium-mobilizing agonists to generate biologically active inositol phosphates in pancreatic acinar cells.

Intracellular calcium levels do not change during contact-mediated collapse of chick DRG growth cone structure

When the growth cone of a chick dorsal root ganglion (DRG) neurite contacts the neurite of a chick retinal ganglion cell in vitro, the growth cone typically responds by withdrawing its lamellipodia and filopodia and collapsing. We have used the fluorescent calcium indicator dye fura-2 and digital imaging microscopy to measure calcium levels within DRG growth cones and to determine whether changes in calcium levels are responsible for the collapse of growth cone morphology when a DRG growth cone contacts a retinal ganglion cell neurite. Calcium levels within DRG growth cones were stable during neurite outgrowth. Calcium was typically distributed homogeneously throughout the growth cone, though occasionally gradients of free calcium were present. When calcium gradients were observed, calcium levels appeared higher in the active veil regions than in the central core region. Calcium levels in DRG growth cones appeared to remain stable during the period of contact-mediated growth cone collapse. Low concentrations of the calcium ionophore ionomycin increased calcium levels two- to threefold without having any observable morphological effects on DRG growth cones. Likewise, depolarization with 15 mM KCl caused a transient two- to threefold increase in calcium levels without having any observable morphological effect. These results suggest that changes in calcium levels are not responsible for contact-mediated collapse of growth cone structure. A growth cone collapsing activity has been solubilized from embryonic chick brain (Raper and Kapfhammer, 1990). Application of this material to cultures of DRG neurons caused growth cones to collapse but had no effect on calcium levels within the growth cones. The crude growth cone collapsing activity was not blocked by the presence of cobalt, nickel, lanthanum, nifedipine, or reduced-calcium medium, suggesting that transmembrane calcium fluxes were not required for growth cone collapse. These results suggest that the morphological changes associated with the collapse of growth cone structure can be independent of changes in growth cone calcium levels, and that second messengers other than calcium are likely to be involved in the regulation of many growth cone behaviors.

Cytosolic ionized calcium in human platelets: The influence of collagen and a novel antiplatelet agent

Two phases of calcium mobilization were observed when aequorin-loaded human platelets, suspended in a nominally calcium-free medium containing 0.1 mM EGTA, were stimulated with collagen. The first phase coincided with platelet shape change, and the second phase corresponded to aggregation. On the other hand, only one [Ca2+]i peak was found in systems containing 1.0 mM Ca, or 1.0 or 2.0 mM EGTA. A novel antiplatelet compound α,α′-bis [3-(N,N-diethylcarbamoyl)piperidino]-p-xylene dihydrobromide, inhibited both [Ca2+]i peaks. It is suggested that inhibition of the mobilization of intraplatelet calcium stores as well as the blocking of transmembrane calcium flux may be responsible for the platelet aggregation-inhibitory action of this compound.

Extracellular calcium-dependent regulation of transmembrane calcium fluxes in murine keratinocytes.

Because the level of extracellular Ca2+ is an important stimulus for differentiation of epidermal cells in vitro, we characterized the extracellular Ca(2+)-dependent transmembrane Ca2+ fluxes in BALB/MK mouse keratinocytes. Increasing levels of extracellular Ca2+, ranging from 0.07 to 1.87 mM, stimulated the rate of 45Ca2+ uptake into these cells 10- to 70-fold and doubled the rate of 45Ca2+ efflux. The divalent cations, Ni2+ and Co2+, were able to block the influx of Ca2+, but dihydropyridines and verapamil were not. Furthermore, 10 to 100 microM of the trivalent cation La3+ induced a dose-dependent 2- to 100-fold increase of Ca2+ uptake, independently of the level of extracellular Ca2+. These observations suggest that keratinocytes possess a cell-surface "Ca(2+)-receptor," activation of which stimulates the influx of 45Ca2+ through a type of voltage-independent, receptor-operated Ca2+ channels. Epidermal growth factor induced an accumulation of 45Ca2+ of a much smaller magnitude than elevations of the level of extracellular Ca2+, without a detectable increase of Ca2+ efflux. Thus, the divergent cellular responses of keratinocytes to EGF and extracellular Ca2+ may be due, in part, to the distinct changes in transmembrane Ca2+ fluxes that these two stimuli generate. Treatment of cells with type beta transforming growth factor led to a gradual 6-fold increase of the Ca(2+)-activated rate of Ca2+ uptake over a period of 4 hours, but reduced the Ca2+ efflux by approximately 50% within 10 minutes. Thus, type beta transforming growth factor apparently stimulates Ca2+ influx indirectly, but may control the differentiation of keratinocytes by direct inhibition of Ca2+ efflux pumps.

Calcium Channel Modulation of α1- and α2-Adrenergic Pressor Responses in Conscious and Anesthetized Dogs

The influence of halothane and isoflurane on alpha-adrenergic-mediated vasoconstriction before and following calcium channel modulation was investigated in chronically instrumented dogs. After ganglionic, cholinergic, and beta-adrenergic blockade, systemic hemodynamic responses following equieffective pressor doses of phenylephrine (0.6 micrograms/kg iv), a selective alpha 1 agonist, and azepexole [B-HT 933] (20 micrograms/kg iv), a selective alpha 2 agonist, were obtained. The calcium channel stimulator Bay k 8644 (0.5 and 1 micrograms.kg-1.min-1) was infused intravenously for 10 min and phenylephrine and azepexole administered at the end of each infusion. On different days, each dog was subsequently anesthetized with equihypotensive concentrations of halothane (1.7%) or isoflurane (2%) in oxygen and the same pharmacologic interventions were repeated in the presence of halothane or isoflurane. Twenty-one experiments (three groups) using seven chronically instrumented dogs were completed. Halothane and isoflurane produced significant (P less than 0.05) attenuation of the increase in arterial pressure after bolus administration of phenylephrine and azepexole. Bay k 8644 augmented the pressor responses mediated by both phenylephrine and azepexole in all three groups. Thus, halothane and isoflurane nonselectively reduced the pressor response to both alpha 1- and alpha 2-adrenergic receptor stimulation and this was probably not mediated by inhibition of transmembrane calcium flux through dihydropyridine sensitive channels.

Exposure of human neutrophils to exogenous nucleotides causes elevation in intracellular calcium, transmembrane calcium fluxes, and an alteration of a cytosolic factor resulting in enhanced superoxide production in response to FMLP and arachidonic acid

Exposure of human neutrophils to micromolar concentrations of both hydrolyzable and nonhydrolyzable purine nucleotides caused the generation of transient rises in intracellular calcium (Ca2+), Ca2+ fluxes across the membrane, and primed the cells for enhanced production of superoxide (O2-) when subsequently exposed to agonists such as FMLP and arachidonic acid. The neutrophils were most sensitive to adenosine triphosphate (ATP) and ATP-gamma-S, which produced Ca2+ transients and enhanced O2- production at concentrations as low as 1 to 5 mumol/L, with a doubling of O2- generation at 25 to 50 mumol/L. Adenosine diphosphate (ADP), guanosine triphosphate (GTP), and 5′- adenylylimidodiphosphate (AMP-PNP) required approximately 10-fold higher concentrations to cause similar effects. Adenosine did not cause Ca2+ fluxes or a Ca2+ transient and was inhibitory of O2- production. There was a strong correlation between a nucleotide's ability to generate a Ca2+ response and its ability to enhance O2- generation. Nitrogen cavitation and subcellular fractionation of the neutrophils after a brief exposure to ATP, ATP-gamma-S, and AMP-PNP revealed that the enhanced O2- generating capacity was stable and detectable in a cell-free assay system. By combining variously treated cytosolic and membrane fractions, it was found that the enhanced O2- production was attributable to a modification of a component(s) of the cytosol.

Exposure of Human Neutrophils to Exogenous Nucleotides Causes Elevation in Intracellular Calcium, Transmembrane Calcium Fluxes, and an Alteration of a Cytosolic Factor Resulting in Enhanced Superoxide Production in Response to FMLP and Arachidonic Acid

Exposure of human neutrophils to micromolar concentrations of both hydrolyzable and nonhydrolyzable purine nucleotides caused the generation of transient rises in intracellular calcium (Ca2+), Ca2+ fluxes across the membrane, and primed the cells for enhanced production of superoxide (O2-) when subsequently exposed to agonists such as FMLP and arachidonic acid. The neutrophils were most sensitive to adenosine triphosphate (ATP) and ATP-gamma-S, which produced Ca2+ transients and enhanced O2- production at concentrations as low as 1 to 5 mumol/L, with a doubling of O2- generation at 25 to 50 mumol/L. Adenosine diphosphate (ADP), guanosine triphosphate (GTP), and 5'-adenylylimidodiphosphate (AMP-PNP) required approximately 10-fold higher concentrations to cause similar effects. Adenosine did not cause Ca2+ fluxes or a Ca2+ transient and was inhibitory of O2- production. There was a strong correlation between a nucleotide's ability to generate a Ca2+ response and its ability to enhance O2- generation. Nitrogen cavitation and subcellular fractionation of the neutrophils after a brief exposure to ATP, ATP-gamma-S, and AMP-PNP revealed that the enhanced O2- generating capacity was stable and detectable in a cell-free assay system. By combining variously treated cytosolic and membrane fractions, it was found that the enhanced O2- production was attributable to a modification of a component(s) of the cytosol.

Tubuloglomerular feedback responses during peritubular infusions of calcium channel blockers

Experiments were performed in pentobarbital-anesthetized rats to evaluate the dependence of the effector limb of the tubuloglomerular feedback mechanism on transmembrane calcium flux through potential-operated calcium channels. Peritubular capillary infusions of the calcium channel blockers, verapamil and nifedipine, were used to achieve high intrarenal levels without influencing arterial blood pressure. Proximal tubule stop-flow pressure (SFP) and single-nephron glomerular filtration rate (SNGFR) tubuloglomerular feedback responses were obtained during control conditions and during simultaneous peritubular capillary infusion with an isotonic saline solution containing either verapamil or nifedipine. Infusion of either 10(-3) M verapamil or 10(-3) M nifedipine, at a rate of 20 nl/min, increased resting SFP (measured during conditions of zero distal volume delivery) and markedly attenuated both the SFP and SNGFR feedback responses to a late proximal perfusion rate of 30 nl/min. Infusion of verapamil (10(-3) M) also increased the slope of the relationship between SFP and renal arterial perfusion pressure between 80 and 120 mmHg (0.43 +/- 0.03 vs 0.24 +/- 0.02, P less than 0.001, n = 10). These findings support the hypothesis that the preglomerular contractile elements responsive to signals from the macula densa cells are activated by calcium influx through potential-operated calcium channels. Furthermore, the preglomerular contractile elements sensitive to calcium channel blockers can dilate further even when orthograde flow to a single macula densa segment is interrupted.

Interaction of intracellular ion buffering with transmembrane-coupled ion transport.

The role of the Na/Ca exchanger in the control of cellular excitability and tension development is a subject of current interest in cardiac physiology. It has been suggested that this coupled transporter is responsible for rapid changes in intracellular calcium activity during single beats, generation of plateau currents, which control action potential duration, and control of intracellular sodium during Na/K pump suppression, which may occur during terminal states of ischemia. The actual behavior of this exchanger is likely to be complex for several reasons. First, the exchanger transports two ionic species and thus its instantaneous flux rate depends on both intracellular sodium and calcium activity. Secondly, the alteration in intracellular calcium activity, which is caused by a given transmembrane calcium flux, and which controls the subsequent exchanger rate, is a complex function of available intracellular calcium buffering. The buffers convert the ongoing transmembrane calcium fluxes into changes in activity that are a small and variable fraction of the change in total calcium concentration. Using a number of simple assumptions, we model changes in intracellular calcium and sodium concentration under the influence of Na/Ca exchange, Na/K ATPase and Ca-ATPase pumps, and passive sodium and calcium currents during periods of suppression and reactivation of the Na/K ATPase pump. The goal is to see whether and to what extent general notions of the role of the Na/Ca exchanger used in planning and interpreting experimental studies are consistent with its function as derived from current mechanistic assumptions about the exchanger. We find, for example, that based on even very high estimates of intracellular calcium buffering, it is unlikely that Na/Ca exchange alone can control intracellular sodium during prolonged Na/K pump blockade. It is also shown that Na/Ca exchange can contaminate measurements of Na/K pump currents under a variety of experimental conditions. The way in which these and other functions are affected by the dissociation constants and total capacity of the intracellular calcium buffers are also explored in detail.

Bile salt potentiates the action of capsaicin on sensory neurones of guinea-pig ileum

In the isolated guinea-pig ileum, exposure to the sensory stimulant drug capsaicin (1 microM) produced a contraction thought to involve substance P(SP) release from sensory nerves. Bile salt, sodium deoxycholate, potentiated the capsaicin-induced contraction in a concentration-dependent (0.03-10 microM) manner, without influencing contractions produced by exogenous SP or by electrical stimulation of efferent nerves. The bile salt-induced potentiation of the capsaicin response was not modified by hexamethonium or indomethacin. It was, however, abolished by concomitant incubation with Ruthenium Red, which was reported to block transmembrane calcium fluxes and then suppress the SP release from the capsaicin-sensitive sensory nerve terminals. We propose that bile salt, as a calcium ionophore, could activate or sensitize the action of capsaicin on the peripheral terminals of sensory nerves.

Calcium in phase control of the Bulla circadian pacemaker

The circadian pacemaker in the retina of the eye of the marine snail Bulla gouldiana was examined using the whole eye in vitro preparation. Phase-response curves were generated to 6-h pulses of a low calcium EGTA solution and to a hyperpolarizing low potassium-low sodium solution. Both treatments yielded similar phase response curves with phase delays in the late subjective night/early subjective day and phase advances in the late subjective day. The similarity of the phase response curves to hyperpolarizing and low calcium solutions and the absence of additivity when both treatments are combined raises the possibility that both treatments affect the underlying pacemaker through a common mechanism. The persistence of phase shifts to low calcium pulses delivered in the presence of depolarizing light suggests that hyperpolarization is not required for low calcium phase shifting. However, it is possible that both treatments act by reducing a transmembrane calcium flux which is postulated to result from the periodic depolarization of the pacemaker cell membrane during the subjective day. Since a transmembrane calcium flux is known to be essential for both light and depolarization-induced phase shifting, we discuss the hypothesis that calcium fluxes play a pivotal role in the entrainment pathway of the circadian pacemaker.

The Effects of Calcium Channel Blockers on Blood Fluidity

Although vasodilation, direct cardiac actions, or both represent the main properties of calcium channel blockers, there are further pharmacologic effects that may be therapeutically relevant. For example, hemorrheological effects, which have been demonstrated for a variety of calcium antagonists, have received relatively little attention to date. Hemorrheology describes the mechanics of blood and its components. It is of particular interest in the context of cardiovascular disease, as it has been shown that under certain conditions (reduced pump function, impaired vasomotor reserve), parameters of blood fluidity may be crucial for tissue perfusion. Whole-blood viscosity is the dominating factor in large arteries. For geometrical reasons, plasma viscosity and the rheological properties of blood cells may become of paramount importance at the microcirculatory level. In ischemic states, erythrocytes may be depleted of ATP, which they need for maintenance of normal shape and for transformation. This results in rigidification of the red blood cell and hindrance of its passage in the microcirculatory bed. Hence, blood flow deteriorates with the consequence of further unfavorable changes of the "milieu interieur," leading to the induction of a vicious cycle. Although effects on several hemorrheological parameters, for example, whole-blood viscosity, plasma viscosity, and red cell aggregation, can be demonstrated for various calcium channel blockers, the main rheological effects of these compounds are believed to consist in the improvement of erythrocyte deformability. When the ATP-dependent calcium pump is impaired in ischemia, calcium channel blockers may inhibit the slow inward transmembrane calcium flux and prevent the accumulation of intracellular calcium.

Ruthenium red antagonism of the effect of capsaicin on the motility of the isolated guinea-pig ileum

Ruthenium red (1 μM), an inorganic dye which blocks transmembrane calcium (Ca) fluxes in neural tissues, selectively reduced the capsaicin (1 μM)-induced contraction of the guinea-pig ileum and protected the sensory fibers from capsaicin-induced desensitization. The ruthenium red (0.5–1 μM) antagonism of capsaicin-induced inhibition of responses to mesenteric nerve stimulation or field stimulation in the isolated guinea-pig ileum was an example of a similar antagonism of the effect of capsaicin. In view of the known action of ruthenium red on the depolarization-coupled entry of Ca into synaptosomes and the release of transmitter, our results support the proposal that ruthenium red could antagonize the action of capsaicin on the peripheral terminals of sensory nerves by a similar mechanism, thereby suppressing transmitter release and preventing the establishment of desensitization.

Aminoglycoside antibiotics impair calcium entry but not viability and motility in isolated cochlear outer hair cells.

Cochlear outer hair cells have been well established as primary targets of the ototoxic actions of aminoglycoside antibiotics. These cells, isolated from the guinea pig cochlea and maintained in short-term culture, were used as a model for evaluating the acute effects of gentamicin on cell viability, depolarization-induced transmembrane calcium flux, and depolarization-induced motile responses. On the basis of morphology and fluorochromasia, the presence of extracellular gentamicin as high as 5 mM did not affect the viability of the cells for up to 6 hr, the longest time tested. Viable cells showed binding of fluorescently tagged gentamicin to their base but excluded the drug from their cytoplasm. In response to [K+]-depolarization, intracellular calcium levels (monitored with the fluorescent calcium-sensitive dye fluo-3) increased from a resting value of 218 +/- 102 nM to 2,018 +/- 1,077 nM concomitant with a cell shortening of 0.7% +/- 1.3%. The depolarization-induced calcium increase was apparently caused by calcium entry into the cell as it was inhibited by the calcium-channel blocker methoxyverapamil and prevented in the absence of extracellular calcium. Both gentamicin and neomycin blocked the [K+]-induced calcium increase at an IC50 of 50 microM. Despite the inhibition of calcium entry the ability of the outer hair cells to shorten under [K+]-depolarization was not impaired; in fact, cell shortening was even more pronounced in the absence of calcium influx (2.6% +/- 1.4%). This argues effectively against the existence of a calcium-dependent actomyosin-mediated component in [K+]-induced shape changes.(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulation of Voltage-Dependent Contractions by Calcium Channel Activator Bay K 8644 in the Human Upper Urinary Tract in Vitro

The effects of calcium agonist Bay K 8644 on mechanical activity were studied in isolated preparations of the human upper urinary tract. Bay K 8644 increased the amplitude of phasic-rhythmic contractions in calyceal segments in a concentration-dependent way. In inactive and unstimulated ureteral muscle strips, Bay K 8644 did not induce contractions. After depolarization of ureteral segments with an extracellular potassium concentration of 48mmol./l., Bay K 8644 produced a concentration-dependent increase of contractile force and enhanced phasic-rhythmic activity. The EC50 of the drug was 7.23 × 108mol./l. The potassium and calcium concentration-response-curves were shifted to the left and the maximum force development was increased. Tonic contractions induced by norepinephrine in calyceal and pelvic segments were not affected by Bay K 8644, but the tendency for phasic-rhythmic activity was increased. In contrast to calcium-antagonistic dihydropyridines like nifedipine, the dihydropyridine derivative Bay K 8644 displayed completely opposite effects, which are obviously limited to voltage-induced activation. These observations can be explained by assuming that these agents act at sites that are components or are associated with voltage-controlled calcium channels. Occupation of these sites may either increase (Bay K 8644) or decrease (nifedipine) the transmembrane calcium flux into the cell.

Stretch-Inactivated Ion Channels Coexist with Stretch-Activated Ion Channels

Stretch-activated ion channels of animal, plant, bacterial, and fungal cells are implicated in mechanotransduction and osmoregulation. A new class of channel has now been described that is stretch-inactivated. These channels occur in neurons, where they coexist with stretch-activated channels. Both channels are potassium selective. The differing stretch sensitivities of the two channels minimize potassium conductance over an intermediate range of tension, with the consequence that, over this same range, voltage-gated calcium channels are most readily opened. Thus, by setting the relation between membrane tension and transmembrane calcium fluxes, stretch-sensitive potassium channels may participate in the control of calcium-dependent motility in differentiating, regenerating, or migrating neurons.