In heart, rapid and transient changes in intracellular [Ca2+] are fundamental to cell function. Rapid Ca2+ release from the sarcoplasmic reticulum through ryanodine receptor (RyR) channels controls contractility. The RyR channel, however, is not the only intracellular Ca2+ release channel present in isolated heart cells. Like all mammalian cells, ventricular cardiac myocytes contain a significant number of IP3Rs. The IP3Rs are intracellular Ca2+ release channels that are homologous to the RyR channel. The RyR and IP3R have similar permeation properties but are differentially regulated by Ca2+ and IP3. The RyR and IP3R mediated intracellular Ca2+ signaling pathways must operate simultaneously in the same cell. How these pathways can operate simultaneous and maintain their individual signal fidelity is not yet clear. In this proposal, single channel function of cardiac RyR and IP3R Ca2+ release channels will be defined in planar bilayers. The kinetics of single RyR and IP3R channels as they respond to fast, calibrated Ca2+ and IP3 stimuli will measured. Fast ligand stimuli will be generated by flash photolysis. The possibility that phosphorylation impacts the differential regulation of the two types of channels will also be investigated. The overall goal is to establish key determinants of cardiac RyR and IP3R function at the single channel level. This represents an important step (i.e. defining/contrasting RyR and IP3R regulatory mechanisms) in understanding the complexity of local intracellular Ca2+ signaling in the heart.
Aim number1: Determine the location, identify and single channel function of IP3R channels in isolated ventricular myocytes.
Aim number 2: Define the identity and kinetics of the mechanisms that """"""""turn-on"""""""" and """"""""turn-off"""""""" single RyR and IP3R release channels.
Aim number 3: Determine how PKA and PKC dependent phosphorylation impacts function of single RyR and IP3R channels.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL064210-02
Application #
6351615
Study Section
Cardiovascular and Pulmonary Research A Study Section (CVA)
Program Officer
Reinlib, Leslie
Project Start
2000-02-14
Project End
2004-01-31
Budget Start
2001-02-01
Budget End
2002-01-31
Support Year
2
Fiscal Year
2001
Total Cost
$248,838
Indirect Cost
Name
Loyola University Chicago
Department
Physiology
Type
Schools of Dentistry
DUNS #
791277940
City
Maywood
State
IL
Country
United States
Zip Code
60153
Ramos-Franco, Josefina; Gomez, Ana M; Nani, Alma et al. (2010) Ryanodol action on calcium sparks in ventricular myocytes. Pflugers Arch 460:767-76
Fill, Michael; Ramos, Jorge (2004) Calcium regulation of single cardiac ryanodine receptor channels. J Muscle Res Cell Motil 25:603-4
Zoghbi, M E; Copello, J A; Villalba-Galea, C A et al. (2004) Differential Ca2+ and Sr2+ regulation of intracellular divalent cations release in ventricular myocytes. Cell Calcium 36:119-34
Ramos, Jorge; Jung, Wonyong; Ramos-Franco, Josefina et al. (2003) Single channel function of inositol 1,4,5-trisphosphate receptor type-1 and -2 isoform domain-swap chimeras. J Gen Physiol 121:399-411
Kettlun, Claudia; Gonzalez, Adom; Rios, Eduardo et al. (2003) Unitary Ca2+ current through mammalian cardiac and amphibian skeletal muscle ryanodine receptor Channels under near-physiological ionic conditions. J Gen Physiol 122:407-17