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Influx Pathway (influx + pathway)
Selected AbstractsDynamics of Ca2+ and Na+ in the dendrites of mouse cerebellar Purkinje cells evoked by parallel fibre stimulationEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2003Akinori Kuruma Abstract Ca2+ and Na+ play important roles in neurons, such as in synaptic plasticity. Their concentrations in neurons change dynamically in response to synaptic inputs, but their kinetics have not been compared directly. Here, we show the mechanisms and dynamics of Ca2+ and Na+ transients by simultaneous monitoring in Purkinje cell dendrites in mouse cerebellar slices. High frequency parallel fibre stimulation (50 Hz, 3,50-times) depolarized Purkinje cells, and Ca2+ transients were observed at the anatomically expected sites. The magnitude of the Ca2+ transients increased linearly with increasing numbers of parallel fibre inputs. With 50 stimuli, Ca2+ transients lasted for seconds, and the peak [Ca2+] reached ,100 µm, which was much higher than that reported previously, although it was still confined to a part of the dendrite. In contrast, Na+ transients were sustained for tens of seconds and diffused away from the stimulated site. Pharmacological interventions revealed that Na+ influx through ,-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and Ca2+ influx through P-type Ca channels were essential players, that AMPA receptors did not operate as a Ca2+ influx pathway and that Ca2+ release from intracellular stores through inositol trisphosphate receptors or ryanodine receptors did not contribute greatly to the large Ca2+ transients. [source] Membrane Hyperpolarization Is Not Required for Sustained Muscarinic Agonist-Induced Increases in Intracellular Ca2+ in Arteriolar Endothelial CellsMICROCIRCULATION, Issue 2 2005KENNETH D. COHEN ABSTRACT Objective: Hyperpolarization modulates Ca2+ influx during agonist stimulation in many endothelial cells, but the effects of hyperpolarization on Ca2+ influx in freshly isolated arteriolar endothelial cells are unknown. Therefore, the purpose of the present study was to characterize agonist-induced Ca2+ transients in freshly isolated arteriolar endothelial cells and to test the hypothesis that membrane hyperpolarization augments agonist-induced Ca2+ influx into these cells. Methods: Arterioles were removed from hamster cremaster muscles and arteriolar endothelial cells were enzymatically isolated. Endothelial cells were loaded with Fura 2-AM and the Fura 2 ratio measured photometrically as an index of intracellular Ca2+. The cells were then stimulated with the muscarinic, cholinergic agonist, methacholine, and the resulting Ca2+ transients were measured. Results: Methacholine (1 , M) increased the endothelial cell Fura 2 ratio from a baseline of 0.81 ± 0.02 to an initial peak of 1.17 ± 0.05 (n = 17) followed by a sustained plateau of 1.12 ± 0.07. The plateau phase of the Ca2+ transient was inhibited by removal of extracellular Ca2+ (n = 12, p < .05), or the nonselective cation channel blockers Gd3+ (30 , M; n = 7, p < .05) or La3+ (50 , M; n = 7, p < .05) without significant effect on the baseline or peak (p > .05). The initial peak of methacholine-induced Ca2+ transients was inhibited by the IP3 -receptor antagonist xestospongin D (10 , M, n = 5, p < .05). The methacholine-induced Ca2+ transients were accompanied by endothelial cell hyperpolarization of approximately 14,18 mV, as assessed by experiments using the potentiometric dye, di-8-ANEPPS as well as by patch-clamp experiments. However, inhibition of hyperpolarization by blockade of Ca2+ -activated K+ channels with charybdotoxin (100 nM) and apamin (100 nM) (n = 5), or exposure of endothelial cells to 80 or 145 mM KCl (both n = 7) had no effect on the plateau phase of methacholine-induced Ca2+ transients (p > .05). Conclusions: Freshly isolated arteriolar endothelial cells display agonist-induced Ca2+ transients. For the muscarinic agonist, methacholine, these Ca2+ transients result from release of Ca2+ from intracellular stores through IP3 receptors, followed by sustained influx of extracellular Ca2+. While these changes in intracellular Ca2+ are associated with endothelial cell hyperpolarization, the methacholine-induced, sustained increase in intracellular Ca2+ appears to be independent from this change in membrane potential. These data suggest that arteriolar endothelial cells may possess a novel Ca2+ influx pathway, or that the relationship between intracellular Ca2+ and Ca2+ influx is more complex than that observed in other endothelial cells. [source] Over-expression of OsAGAP, an ARF-GAP, interferes with auxin influx, vesicle trafficking and root developmentTHE PLANT JOURNAL, Issue 4 2006Xiaolei Zhuang Summary Development and organogenesis in both dicot and monocot plants are highly dependent on polar auxin transport (PAT), which requires the proper asymmetric localization of both auxin influx and efflux carriers. In the model dicot plant Arabidopsis thaliana, the trafficking and localization of auxin efflux facilitators such as PIN-FORMED1 (PIN1) are mediated by GNOM, a guanine-nucleotide exchange factor (GEF) for the ADP-ribosylation factor (ARF) family of small GTPases, but molecular regulators of the auxin influx facilitators remain unknown. Here, we show that over-expression of OsAGAP, an ARF-GTPase-activating protein (ARF-GAP) in rice, impaired PAT and interfered with both primary and lateral root development. The lateral root phenotype could be rescued by the membrane-permeable auxin 1-naphthyl acetic acid, but not by indole 3-acetic acid (IAA) or by 2,4-dichloro-phenoxyacetic acid, which require influx facilitators to enter the cells. OsAGAP-over-expressing plants had alterations in vesicle trafficking and localization of the presumptive A. thaliana auxin-influx carrier AUX1, but not in the localization of the auxin efflux facilitators. Together, our data suggest that OsAGAP has a specific role in regulating vesicle trafficking pathways such as the auxin influx pathway, which in turn controls auxin-dependent root growth in plants. [source] Electrophysiological effects of endothelin-1 and their relationship to contraction in rat renal arterial smooth muscleBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2000Luisa C Betts The electophysiological effects of endothelin-1 (ET-1) and their relationship to contraction remain unclear in the renal circulation. Using endotheliumdenuded arteries from the main branch of the renal artery proximal to the kidney of the rat, we have examined its effects on tension and conducted parallel patch-clamp measurements using freshly isolated smooth muscle cells from this tissue. Pharmacological experiments revealed that ET-1 produced constriction of renal arteries dependent on the influx of extracellular Ca2+, mediated solely through ETA receptor stimulation. Current-clamp experiments revealed that renal arterial myocytes had a resting membrane potential of ,32 mV, with the majority of cells exhibiting spontaneous transient hyperpolarizations (STHPs). Application of ET-1 produced depolarization and in those cells exhibiting STHPs, either caused their inhibition or made them occur regularly. Under voltage-clamp conditions cells were observed to exhibit spontaneous transient outward currents (STOCs) inhibited by iberiotoxin. Application of voltage-ramps revealed an outward current activated at ,,30 mV, sensitive to both 4-AP and TEA. Taken together these results suggest that renal arterial myocytes possess both delayed rectifying K+ (KV) and Ca2+ -activated K+ (BKCa) channels. Under voltage-clamp, ET-1 attenuated the outward current and reduced the magnitude and incidence of STOCs: effects mediated solely as a consequence of ETA receptor stimulation. Thus, in conclusion, activation of ETA receptors by ET-1 causes inhibition of KV and BKCa channel activity, which could promote and/or maintain membrane depolarization. This effect is likely to favour L-type Ca2+ channel activity providing an influx pathway for extracellular Ca2+ essential for contraction. British Journal of Pharmacology (2000) 130, 787,796; doi:10.1038/sj.bjp.0703377 [source] | |||||||||||||