Oral administration of sodium pyrithione (NaP) causes hindlimb weakness in rodents, but not in primates. Previous work using Aplysia neurons has demonstrated that NaP produces a persistent influx of Ca2+ ions across the plasma membrane. To determine whether this also occurs in mammalian neurons and whether this could underlie the inter-species difference between rodents and primates, we have tested the effects of NaP on intracellular Ca2+ levels ([Ca2+]i) in rat and monkey motor neurons in vitro. Motor neurons present in spinal cord slices from rhesus monkey embryos (E37 and 56) and from rat E16 were dissected and cultured on glass coverslips. Following 2 weeks (rhesus) or 2-3 days (rat) in culture, neurons were loaded with fura-PE3/AM, and examined for [Ca2+]i changes in response to NaP. Rhesus motor neurons were identified by immunostaining for Islet-1 (MN specific antigen) and neuron specific enolase (NSE). Motor neurons from both species exhibited dose-dependent NaP-evoked increases in [Ca2+]i However, the dose-response curve for the Rhesus motor neurons was significantly shifted to the right of the rat dose-response curve, whereas the overall amplitude of the Ca2+ rise was similar in both species. As shown previously for the Aplysia neurons, the action of NaP is attenuated by SKF 96365, an inhibitor of store-operated calcium entry. In contrast the action of NaP is unaffected by nifedipine and tetrodotoxin, blockers of voltage-dependent Ca2+ and Na+ channels, respectively, or by ouabain, an inhibitor of the plasma membrane Na+/K+ ATPase. Our results indicate that the NaP-induced increase in [Ca2+]i is conserved across species and suggest that the toxicological sensitivity of rodent over primate to pyrithione could be due to the enhanced sensitivity of rodent motor neurons to NaP-evoked intracellular Ca2+ elevation. 相似文献
2-Methoxyestradiol, an endogenous metabolite of 17β-estradiol, is known to have antitumor and antiangiogenic actions. The
effects of 2-methoxyestradiol on ionic currents were investigated in an endothelial cell line (HUV-EC-C) originally derived
from human umbilical vein. In the whole-cell patch-clamp configuration, 2-methoxyestradiol (0.3–30 μm) reversibly suppressed the amplitude of K+ outward currents. The IC50 value of the 2-methoxyestradiol-induced decrease in outward current was 3 μm. Evans blue (30 μm) or niflumic acid (30 μm), but not diazoxide (30 μm), reversed the 2-methoxyestradiol-induced decrease in outward current. In the inside-out configuration, application of 2-methoxyestradiol
(3 μm) to the bath did not modify the single-channel conductance of large-conductance Ca2+-activated K+ (BKCa) channels; however, it did suppress the channel activity. 2-Methoxyestradiol (3 μm) produced a shift in the activation curve of BKCa channels to more positive potentials. Kinetic studies showed that the 2-methoxyestradiol-induced inhibition of BKCa channels is primarily mediated by a decrease in the number of long-lived openings. 2-Methoxyestradiol-induced inhibition
of the channel activity was potentiated by membrane stretch. In contrast, neither 17β-estradiol (10 μm) nor estriol (10 μm) affected BKCa channel activity, whereas 2-hydroxyestradiol (10 μm) slightly suppressed it. Under current-clamp condition, 2-methoxyestradiol (10 μm) caused membrane depolarization and Evans blue (30 μm) reversed 2-methoxyestradiol-induced depolarization. The present study provides evidence that 2-methoxyestradiol can suppress
the activity of BKCa channels in endothelial cells. These effects of 2-methoxyestradiol on ionic currents may contribute to its effects on functional
activity of endothelial cells.
Received: 27 November 2000/Revised: 13 April 2001 相似文献
A translational lacZ fusion of the Bacillus subtilis mscL gene that encodes the mechanosensitive channel of large conductance (MscL) was expressed at significant levels during log phase growth of B. subtilis, and the level of mscL–lacZ expression was increased 1.5-fold by growth in medium with high salt (1 M NaCl). However, in growth media with either low or high salt, mscL–lacZ expression fell drastically beginning in the late log phase of growth, and fell to even lower levels during sporulation, although a significant amount of β-galactosidase from mscL to lacZ was accumulated in the developing spore. Deletion of mscL had no effect on B. subtilis growth, sporulation or subsequent spore germination. The ΔmscL strain also grew as well as the wild-type parental strain in medium with 1.2 M NaCl. While log phase wild-type cells grown with 1.2 M NaCl survived a rapid 0.9 M osmotic downshift, log phase ΔmscL cells rapidly lost viability and lysed when subjected to this same osmotic downshift. However, by the early stationary phase of growth, ΔmscL cells had become resistant to a 0.9 M osmotic downshift. 相似文献
A1 adenosine receptors (ARs) reduce, and A2ARs increase intraocular pressure, partly by differentially altering resistance to aqueous humor outflow. It is unknown whether the opposing effects of A1AR and A2AR agonists are mediated at different outflow-pathway cell targets or by opposing actions on a single cell target. We tested whether a major outflow-pathway cell, the trabecular meshwork (TM) cell might constitute the primary AR-agonist target and respond differentially to A1, A2A and A3AR agonists. Receptor activation in human TM cells was identified by applying subtype-selective AR agonists: CPA and ADAC for A1ARs, CGS 21680 and DPMA for A2AARs, and Cl-IB-MECA and IB-MECA for A3ARs. Stimulation of A1, A2A and A3ARs elevated Ca2+, measured with fura-2. Whole-cell patch clamping indicated that AR agonists activated ion channels non-uniformly, possibly reflecting variability in magnitude of agonist-triggered second-messenger responses. A1, A2A and A3AR agonists all reduced volume, determined by calcein cell imaging. The endogenous source of adenosine delivery to the outflow pathway could be the TM cells since these cells were stimulated to release ATP by hypotonic perfusion. We conclude that: (1) TM cells express functional A1, A2A and A3ARs; and (2) the reported differential effects of AR agonists on aqueous humor outflow are not mediated by differential actions on TM-cell Ca2+ and volume, but likely by actions on separate cell targets.
Reprint requests should be addressed to: Dr. Mortimer M. Civan, Dept. of Physiology, University of Pennsylvania, Richards Building, Philadelphia, PA 19104-6085. [Tel.: (215)-898-8773; Fax: (215)-573-5851] 相似文献
Calcium (Ca2+) is a ubiquitous second messenger that regulates a plethora of physiological functions. Deregulation of calcium homeostasis has been reported in a wide variety of pathological conditions including cardiovascular disorders, cancer and neurodegenerative diseases. One of the most ubiquitous pathways involved in regulated Ca2+ influx into cells is the store-operated Ca2+ entry (SOCE) pathway. In 2006, Orai1 was identified as the channel protein that mediates SOCE in immune cells. Orai1 has two mammalian homologs, Orai2 and Orai3. Although Orai1 has been the most widely studied Orai isoform, Orai3 has recently received significant attention. Under native conditions, Orai3 was demonstrated to be an important component of store-independent arachidonate-regulated Ca2+ (ARC) entry in HEK293 cells, and more recently of a store-independent leukotrieneC4-regulated Ca2+ (LRC) entry pathway in vascular smooth muscle cells. Recent studies have shown upregulation of Orai3 in estrogen receptor-expressing breast cancers and a critical role for Orai3 in breast cancer development in immune-compromised mice. Orai3 upregulation was also shown to contribute to vascular smooth muscle remodeling and neointimal hyperplasia caused by vascular injury. Furthermore, Orai3 has been shown to contribute to proliferation of effector T-lymphocytes under oxidative stress. In this review, we will discuss the role of Orai3 in reported pathophysiological conditions and will contribute ideas on the potential role of Orai3 in native Ca2+ signaling pathways and human disease. 相似文献
In neurons of the rat dorsal root ganglia (DRG), using a patch-clamp technique in the whole-cell configuration, we studied
the characteristics of calcium channels activated by depletion of the ryanodine-sensitive calcium stores of the endoplasmic
reticulum. Current-voltage (I-V) relationships of these store-operated calcium channels were obtained by subtraction of the
integral I-V characteristics after application of caffeine from the integral I-V characteristics of calcium channels in the
control. Currents through store-operated calcium channels could be induced by application of a series of hyperpolarization
current pulses to the cell under conditions of replacement of a calcium-free solution containing caffeine by a caffeine-free
solution containing 2 mM Ca2+. In this case, the following two main conditions were abserved: Voltage-operated calcium channels were inactivated, while
a gradient of the electrochemical potential for calcium ions was increased, which made easier passing of these currents through
store-operated calcium channels. Therefore, we found that in DRG neurons, despite the presence of great numbers of both voltage-operated
and receptor-dependent calcium channels, one more mechanism underlying the entry of calcium through store-operated channels
does exist.
Neirofiziologiya/Neurophysiology, Vol. 39, No. 3, pp. 195–200, May–June, 2007. 相似文献
Summary The present studies examined some of the properties of Cl– channels in renal outer medullary membrane vesicles incorporated into planar lipid bilayers. The predominant channel was anion selective having aPCl/PK ratio of 10 and a unit conductance of 93 pS in symmetric 320mm KCl. In asymmetric KCl solutions, theI-V relations conformed to the Goldman-Hodgkin-Katz equation. Channel activity was voltage-dependent with a gating charge of unity. This voltage dependence of channel activity may account, at least in part, for the striking voltage dependence of the basolateral membrane Cl– conductance of isolated medullary thick ascending limb segments. The Cl– channels incorporated into the planar bilayers were asymmetrical: thetrans surface was sensitive to changes in ionized Ca2+ concentrations and insensitive to reducing KCl concentrations to 10mm, while thecis side was insensitive to changes in ionized Ca2+ concentrations, but was inactivated by reducing KCl concentrations to 50mm. 相似文献
The mechanical state of the heart feeds back to modify cardiac rate and rhythm. Mechanical stretch of myocardial tissue causes immediate and chronic responses that lead to the common end point of arrhythmia. This review provides a brief summary of the author's personal choice of contributions that she considers have fostered our understanding of the role of mechano-electric feedback in arrhythmogenesis.
Acute mechanical stretch reversibly depolarises the cell membrane and shortens the action potential duration. These electrophysiological changes are related to the activation of mechano-sensitive ion channels. Several different ion channels are involved in the sensing of stretch, among them K+-selective, Cl−-selective, non-selective, and ATP-sensitive K+ channels. Sodium and Ca2+ entering the cells via non-selective ion channels are thought to contribute to the genesis of stretch-induced arrhythmia. Mechano-sensitive channels have been cloned from non-vertebrate and vertebrate species.
Chronic stress on the heart activates gene expression in cardiomyocytes and non-myocytes. The signal transduction involves atrial natriuretic peptides and growth factors that initiate remodelling processes leading to hypertrophy which in turn may contribute to the electrical instability of the heart by increasing the responsiveness of mechano-sensitive channels. Selective block of these channels could provide some new form of treatment of mechanically induced arrhythmias, although at present there are no drugs available with sufficient selectivity. Detailed understanding of how mechanical strain on myocardial cells is translated into channel activation will allow to identify new targets for putative antiarrhythmic drugs. 相似文献
A theoretical model of calcium signaling is presented that simulates oscillations of cytoplasmic calcium concentration ([Ca2+]cyt) in stomatal guard cells under the action of abscisic acid. The model is based on the kinetics of inositol 1,4,5-trisphosphate-sensitive calcium channels of endoplasmic reticulum and cyclic ADP-ribose-sensitive calcium channels of the tonoplast. The operation of two energy-dependent pumps—the Ca2+-ATPase of the endoplasmic reticulum and the Ca2+/H+ antiporter of the tonoplast—is also included in the model. It is shown that the removal of excessive Ca2+ from the cytoplasm by the tonoplast Ca2+/H+ antiporter is the main factor accounting for generation of [Ca2+]cyt oscillations at a wide range of ABA concentrations (0.01–1 M). The long period of [Ca2+]cyt oscillations in plant cells is explained by a slow release from inhibition of inositol 1,4,5-trisphosphate-gated calcium channels. 相似文献