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The use of cytoskeletal inhibitors to determine the role of the cytoskeleton in the activation of hypertonicity-induced currents in xenopus oocytes

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Identifier	Thesis 1561
Author	Balanda, Matthew L.
Title	The use of cytoskeletal inhibitors to determine the role of the cytoskeleton in the activation of hypertonicity-induced currents in xenopus oocytes
Publisher	Central Connecticut State University
Date	1999
Resource Type	Master's Thesis
Notes	Our lab has recently observed a triphasic hypertonicity induced current in stage IV-VI Xenopus laevis oocytes. This current is a calcium-dependant chloride current which is similar to that observed by (Hartzell, 1996). Hartzell, 1996) found that this triphasic chloride current was activated shortly following the injection of inositol (1,4,5)-trisphosphate into Xenopus laevis oocytes. These two findings suggest that the hypertonicity-induced current, Ihi, is activated through a signal transduction cascade involving phospholipase-C. the focus of these experiments was to determine whether or not decreases in cell volume can induce mechanical changes in the cytoskeleton of the oocyte that will initiate this signal transduction cascade. Cells were either pretreated with 33μM nocodazole, 50μM cytochalasin B, 2.5μM taxol, or 0.05% DMSO. Each cell was voltage clamped at -65 mV and ramped to +30mV every 10 seconds throughout the course of the experiment. Ihi peak and plateau currents at -60mV and +10mV were recorded as the cells were perfused with 400 mOsm/kg hypertonic ND96. Cell volume measurements were also recorded. There were only three significant differences between the treatment and control groups. Cells pretreated with 33μM nocodazole shrank significantly more than control cells, p<0.05, when exposed to the hypertonic ringer solution. This same batch of cells also elicited a greater peak shrinkage activated current, Ihipeak, at -60mV, p<0.05, than the control cells. Cells pretreated with 50μM cytochalasin B had a significant reduction in Ihipeak at +10mV when compared to control cells, P<0.05. Microtubules are radically orientated throughout the oocyte (Gard, et al., 1997). The extensive shrinkage in cells pretreated with microtubule disrupters, like nocodazole, would seem to suggest that the arrangement of microtubules is somehow involved in maintenance of cell size and shape during shrinkage. Inhibition of Ihi at +10mV in cytochalasin B treated cells suggests a possible involvement of the cytoskeleton, specifically the microfilaments, in the initiation of the signal transduction cascade that brings about the activation of Ihi. Increased peak shrinkage currents at -60mV in the nocodazole treated cells was not expected. Nevertheless, combined with the other data presented, this could indicate the possibility that changes in cytoskeletal tension might indeed be able to cause conformational changes in one or more of the proteins involved in the activation of the hypertonicity induced current.
Subject	Cytoskeleton Xenopus laevis Cytology
Department	Department of Biological Sciences
Advisor	Martin-Troy, Kathy
Type	Text
Digital Format	application/pdf
Language	eng
OCLC number	713735052
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The use of cytoskeletal inhibitors to determine the role of the cytoskeleton in the activation of hypertonicity-induced currents in xenopus oocytes

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