DEVELOPMENT OF CARDIAC EXCITATION-CONTRACTING COUPLING

描述：（申请人摘要）

DESCRIPTION: (Applicant's Abstract) In adult cardiac muscle, depolarization activates dihydropyridine (DHP) sensitive Ca channels in the transverse tubule (t-tube) membrane. The open DHP Ca channel carries a small Ca flux into the cell. This small Ca flux acts as a trigger Ca signal that activates Ca channels in the sarcoplasmic reticulum (SR). This proposal seeks to examine how the microscopic attributes of the trigger Ca signal govern the activity of single SR Ca release channels and thus generate the local Ca release events essential to normal cardiac function. The SR Ca release channel has been identified as the ryanodine receptor (RyR). Local Ca control of single RyR channels in adult and neonate rat ventricular myocytes is different. Adult RyR channel function is clearly governed by trigger Ca signals rising from the opening of juxtaposed DHP sensitive Ca channels. Neonate RyR channel function depends on a very different set morphological constraints and factors. For example, neonate cells have no t-tubes and lacks the high degree of SR ultrastructural specialization seen in adult. Defining the local Ca control mechanisms that govern adult and neonate RyR function will, 1) increase knowledge concerning development of an intricate intracellular Ca signaling system (i.e. cardiac E-C coupling) and, 2) provide new insights into RyR local Ca control mechanisms in general. The single channel behavior of adult and neonate RyR will be defined, compared, and used to generate comprehensive dynamic Markovian models RyR function (planar bilayer studies in Aim #1). The macro- and microscopic trigger Ca signals generated by the action potential (AP) in adult and neonate acutely dissociated myocytes will be measured (patch-clamp & confocal imaging studies in Aim #2). The response of single adult and neonate RyR channels to these complex trigger Ca waveforms will be predicted and experimentally tested (modeling & bilayer studies in Aim #3). The spatio-temporal attributes of evoked and spontaneous intracellular Ca signals in adult and neonate cells will be defined and correlated to the local Ca control of single adult and neonate RyR channels (modeling & confocal imaging studies, Aim #4).