Laurence S. David, Esperanza Garcia, Stuart M. Cain, Elana Thau, John R. Tyson and Terrance P. Snutch

Laurence S. David

University of British Columbia; Vancouver, BC Canada

Esperanza Garcia

University of British Columbia; Vancouver, BC Canada

Stuart M. Cain

University of British Columbia; Vancouver, BC Canada

Elana Thau

University of British Columbia; Vancouver, BC Canada

John R. Tyson

University of British Columbia; Vancouver, BC Canada

Terrance P. Snutch

Corresponding author: snutch@msl.ubc.ca
University of British Columbia; Vancouver, BC Canada

Low voltage-activated T-type calcium (Ca) channels contribute to the normal development of the heart and are also implicated in pathophysiological states such as cardiac hypertrophy. Functionally distinct T-type Ca channel isoforms can be generated by alternative splicing from each of three different T-type genes (Ca_V3.1, Ca_V3.2,Ca_V3 .3), although it remains to be described whether specific splice variants are associated with developmental states and pathological conditions. We aimed to identify and functionally characterize Ca_V3.2 T-type Ca channel alternatively spliced variants from newborn animals and to compare with adult normotensive and spontaneously hypertensive rats (SHR). DNA sequence analysis of full-length Ca_V3.2 cDNA generated from newborn heart tissue identified ten major regions of alternative splicing, the more common variants of which were analyzed by quantitative real-time PCR (qRT-PCR) and also subject to functional examination by whole-cell patch clamp. The main findings are that: (1) cardiac Ca_V3.2 T-type Ca channels are subject to considerable alternative splicing, (2) there is preferential expression ofCa_V3 .2(-25) splice variant channels in newborn rat heart with a developmental shift in adult heart that results in approximately equal levels of expression of both (+25) and (-25) exon variants, (3) in the adult stage of hypertensive rats there is a both an increase in overallCa_V3 .2 expression and a shift towards expression of Ca_V3.2(+25) containing channels as the predominant form, and (4) alternative splicing confers a variant-specific voltage-dependent facilitation ofCa_V3 .2 channels. We conclude that Ca_V3.2 alternative splicing generates significant T-type Ca channel structural and functional diversity with potential implications relevant to cardiac developmental and pathophysiological states.