UVM Theses and Dissertations
Format:
Print
Author:
Pachuau, Judith L.
Dept./Program:
Biology
Year:
2008
Degree:
PhD
Abstract:
Voltage gated Ca²⁺ channels are the major source of intracellular Ca²⁺ in neurons. Once Ca²⁺ enters the cells, a variety of events can take place including differentiation, proliferation and cell death. Although the physiological functions of voltage-gated Ca²⁺ channels are well established, we have very little understanding of their regulation during development.
The functional expression of T-type Ca²⁺ channels in nodose neurons is upregulated by culture with a cardiac extract. The functional expression of T-type Ca²⁺ channels reaches a maximum effect after 12 hr. This effect is mimicked by hematopoietic cytokines such as CNTF and LIF. The upregulation of T-type Ca²⁺ channels by cardiac extract and cytokines does not seem to involve protein synthesis but instead appears to be a posttranslational event. The stimulatory effect of heart extract and CNTF involves signaling through the Erk MAP kinase and Jak-STAT signaling pathways. Western blot analysis indicates that the Jak inhibitors P6 and AG490 as well as the Erk inhibitor U0126 cause a complete inhibition of the Jak-STAT and Erk signaling pathways evoked by cardiac extract or CNTF. These inhibitors also cause a significant inhibiton of T-type Ca²⁺ channel expression evoked by heart extract and CNTF.
Since the upregulation of T-type Ca²⁺ channels involves a posttranslational event, we looked at the role of the actin cytoskeleton in regulating trafficking of these channels to the membrane. Phalloidin, an actin-stabilizing drug completely blocked the expression of T-type channels without affecting HVA currents. However, latrunculin-A and cytochalasin-D, drugs that prevent actin depolymerization have no effect on channel expression. This suggests that the functional expression of T-type Ca²⁺ channels requires a dynamic actin cytoskeleton. Microtubules have also been implicated in the functional expression of ion channels. However, the disruption of microtubules by nocodazole and colchicine has no effect on T-type Ca²⁺ channel expression. Pharmacological evidence suggests that small GTPases such as ras and arf may be involved in the upregulation of T-type Ca²⁺ channels upon stimulation.
The functional expression of T-type Ca²⁺ channels in nodose neurons is upregulated by culture with a cardiac extract. The functional expression of T-type Ca²⁺ channels reaches a maximum effect after 12 hr. This effect is mimicked by hematopoietic cytokines such as CNTF and LIF. The upregulation of T-type Ca²⁺ channels by cardiac extract and cytokines does not seem to involve protein synthesis but instead appears to be a posttranslational event. The stimulatory effect of heart extract and CNTF involves signaling through the Erk MAP kinase and Jak-STAT signaling pathways. Western blot analysis indicates that the Jak inhibitors P6 and AG490 as well as the Erk inhibitor U0126 cause a complete inhibition of the Jak-STAT and Erk signaling pathways evoked by cardiac extract or CNTF. These inhibitors also cause a significant inhibiton of T-type Ca²⁺ channel expression evoked by heart extract and CNTF.
Since the upregulation of T-type Ca²⁺ channels involves a posttranslational event, we looked at the role of the actin cytoskeleton in regulating trafficking of these channels to the membrane. Phalloidin, an actin-stabilizing drug completely blocked the expression of T-type channels without affecting HVA currents. However, latrunculin-A and cytochalasin-D, drugs that prevent actin depolymerization have no effect on channel expression. This suggests that the functional expression of T-type Ca²⁺ channels requires a dynamic actin cytoskeleton. Microtubules have also been implicated in the functional expression of ion channels. However, the disruption of microtubules by nocodazole and colchicine has no effect on T-type Ca²⁺ channel expression. Pharmacological evidence suggests that small GTPases such as ras and arf may be involved in the upregulation of T-type Ca²⁺ channels upon stimulation.