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Dual regulation of R-type CaV2.3 current by Gq-coupled receptors

Title
Dual regulation of R-type CaV2.3 current by Gq-coupled receptors
Authors
Jeong, Jin Young
DGIST Authors
Jeong, Jin Young; Suh, Byung Chang; Kim, Ho Jeong
Advisor(s)
Suh, Byung Chang
Co-Advisor(s)
Kim, Ho Jeong
Issue Date
2015
Available Date
2015-07-19
Degree Date
2015. 8
Type
Thesis
Keywords
CaV2.3 channelM1 muscarinic receptorphosphatidylinositol 45-bisphosphate (PI(45)P2)
Abstract
Many high voltage-activated Ca2+ channels are modulated by Gq-coupled M1 muscarinic acetylcholine receptors. CaV2.3 currents are known to be increased by M1 receptor activation, and the increase in the CaV2.3 currents is mediated by phosphorylation of CaV2.3 channel via the activation of protein kinase C (PKC). Here, we report that M1 muscarinic receptors can also inhibit CaV2.3 currents when the channels are fully activated by PKC. In the whole-cell configuration of tsA201 cells, phorbol 12-myristate 13-acetate (PMA), a PKC activator, potentiated CaV2.3 currents by ~ 2-fold. We found that after the PMA-induced potentiation of CaV2.3 currents, application of the M1 receptor agonist oxotremorine-M (Oxo-M), decreased the currents by 52%. We examined if the hydrolysis of plasma membrane phosphoinositides (PIs) were involved in the muscarinic suppression of CaV2.3 currents. We used two methods to deplete PI(4,5)P2; voltage-sensing phosphatase (VSP), and rapamycin-induced translocatable pseudojanin (PJ) system. Activation of VSP suppressed CaV2.3 current by 38%. PJ system could directly dephosphorylate 4- and 5-phosphates from both PI(4)P and PI(4,5)P2 the plasma membrane. After the addition of rapamycin CaV2.3 currents were dramatically and irreversibly decreased by 66% compared to the initial level. Taken together, our results suggest that CaV2.3 currents are modulated by M1 receptor in a dual mode; potentiation by PKC activation and suppression by poly-PI depletion. Activation of M1 receptors can solely decrease CaV2.3 currents in the PKC-activated cells. PJ-induced inhibition of CaV2.3 currents demonstrates that poly-PIs are important in the maintenance of CaV2.3 channel activity. ⓒ 2015 DGIST
Table Of Contents
1. Introduction 1 -- 2. Materials and methods 4 -- 2.1 Materials 4 -- 2.2 Cell culture 4 -- 2.3 Transfection 5 -- 2.4 Solution 5 -- 2.5 Chemicals 6 -- 2.6 Current recording 6 -- 2.7 Confocal imaging 6 -- 2.8 Data analysis 7 -- 3. Results 8 -- 3.1 CaV2.3 currents are suppressed as well as stimulated by M1 muscarinic receptor 8 -- 3.2 CaV2.3 currents are decreased by Dr-VSP activation 9 -- 3.3 CaV2.3 currents are decreased by chemically-induced phosphoinositide depletion 9 -- 4. Discussion 12 -- 5. Figure legends 15 -- 6. Figures 19 -- References 32 -- Abstract in Korean 36
URI
http://dgist.dcollection.net/jsp/common/DcLoOrgPer.jsp?sItemId=000002064916
http://hdl.handle.net/20.500.11750/1418
DOI
10.22677/thesis.2064916
Degree
Master
Department
Brain and Cognitive Sciences
University
DGIST
Files:
Collection:
Brain and Cognitive SciencesThesesMaster


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