Parathyroid hormone (PTH) release is regulated by changes in the extracellular (EC) [Ca2+]. High Ca2+ inhibits, and low Ca2+ stimulates PTH secretion. Raising EC Ca2+ produces prompt, sustained increases in intracellular free Ca2+ ([Ca 2+]i) and the Ca2+- mobilizing second-messenger inositol trisphosphate (Insp3)- 1,4,5-Insp3 releases Ca 2+ from intracellular stores in a variety of cells and, in T-cells, can also stimulate Ca2+ influx by opening membrane Ca2+ channels. In sea urchin eggs and lacrimal cells, 1,3,4,5 InsP4, generated from 1,4,5-Insp3 by 1,4,5-InsP3 3-kinase and ATP, acts together with 1,4,5-Insp3 to mediate membrane Ca2+ influx. The role of 1,4,5-Insp3 and 1,3,4,5-Insp4 in the regulation of EC Ca2+- induced increases in [Ca 2+]i has not been addressed nor have the Ca2+ influx mechanisms in parathyroid cells been studied directly by patch-clamp techniques. Our hypothesis is that parathyroid cells express receptors or sensors for Ca 2+ which when activated induce rapid and sustained increases in [Ca2+], due to intracellular Ca2+ mobilization and membrane Ca2+ influx. We propose that 1,4,5-Insp3 mediates the initial release of intracellular Ca2+ and that membrane Ca2+ channels-- gated either by 1,4,5-Insp3 and/or 1,3,4,5-InsP4, by a GTP-binding protein, or by the Ca2+ receptor molecule itself -- are responsible for sustained increases in [Ca2+]i . The goal of the current proposal is to identify and characterize the mechanisms for Ca2+ mobilization in parathyroid cells using electrophysiologic and biochemical approaches. Specifically, we will determine whether high EC Ca2+ induces 1 4,5-InsP3 and 1,3,4,5Insp4 accumulation; whether 1,3,4,5-Insp4 regulates the reuptake of Ca2+ released by 1,4,5-Insp3; and whether 1,4,5-Insp3 production is to essential to Ca2+ mobilization and uptake. We will accomplish the latter by blocking phosphatidyl inositol 4,5-bis P04 (PIP2) hydrolysis and 1,4,5-lnsp3 generation with a phospholipase C inhibitor and a monoclonal antibody which binds to PIP2 and by the 1,4,5-Insp3 analogue 1,4,5-Insp3S, which mobilizes 1,4,5-Insp3-sensitive Ca2+ pools and resists phosphorylation to 1,3,4,5Insp4- By patch-clamping, we will assess whether these cells express voltage-insensitive Ca2+ channels which are regulated by second-messengers or guanyl nucleotides or directly by the Ca2+ receptor molecule. These approaches should provide important insights into the regulation of [Ca2+]i in parathyroid cells and identify Ca2+ influx mechanisms that may be involved in the pathogenesis of PTH hypersecretory states.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK043400-03
Application #
2142987
Study Section
General Medicine B Study Section (GMB)
Project Start
1991-06-01
Project End
1995-03-31
Budget Start
1993-06-01
Budget End
1995-03-31
Support Year
3
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
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