Characterization of the molecular and electrophysiological properties of the T-type calcium channel in human myometrium.
Rises in intracellular calcium are essential for contraction of human myometrial smooth muscle (HMSM) and hence parturition. The T-type calcium channel may play a role in this process. The aim was to investigate the role of the T-type calcium channel in HMSM by characterizing mRNA expression, protein localization, electrophysiological properties and function of the channel subunits Cav3.1(alpha1G), Cav3.2(alpha1H), and Cav3.3(alpha1I). QRT-PCR, immunohistochemistry, electrophysiology and invitro contractility were performed on human myometrial samples from term, preterm, labour and not in labour. QRT-PCR analysis of Cav3.1, Cav3.2 and Cav3.3 demonstrated expression of Cav3.1 and Cav3.2 with no significant change (P>0.05) associated with gestation or labour status. Immunohistochemistry localized Cav3.1 to myometrial and vascular smooth muscle cells whilst Cav3.2 localized to vascular endothelial cells and invading leucocytes. Voltage clamp studies demonstrated a T-type current in 55% of cells. Nickel block of T-type current was voltage sensitive (IC50 of 118.57+/-68.9 microM at -30 mV). Activation and inactivation curves of ICa currents in cells expressing T-type channels overlapped demonstrating steady state window currents at the resting membrane potential of myometrium at term. Current clamp analysis demonstrated that hyperpolarizing pulses to a membrane potential greater than -80 mV elicited rebound calcium spikes that were blocked reversibly by 100 microM nickel. Contractility studies demonstrated a reversible decrease in contraction frequency during application of 100 microM nickel (P<0.05). We conclude that the primary T-type subunit expressed in some MSMCs is Cav3.1. We found that application of 100 microM nickel to spontaneously contracting human myometrium reversibly slows contraction frequency.
Blanks
Andrew M
AM
Clinical Science Research Institute, Division of Clinical Sciences, Warwick Medical School, Coventry CV4 7AL, UK. andrew.blanks@warwick.ac.uk
Zhao
Zheng-Hang
ZH
Shmygol
Anatoly
A
Bru-Mercier
Gilles
G
Astle
Shirley
S
Thornton
Steven
S
D020747:Calcium Channels, T-Type / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling; D002585:Cesarean Section; D005260:Female; D015870:Gene Expression; D005865:Gestational Age; D006801:Humans; D007150:Immunohistochemistry; D007743:Labor, Obstetric / Q000378:metabolism; D008564:Membrane Potentials; D026901:Membrane Transport Proteins / Q000378:metabolism; D009215:Myometrium / Q000187:drug effects / Q000378:metabolism*; D009532:Nickel / Q000494:pharmacology; D036801:Parturition / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D047928:Premature Birth / Q000378:metabolism; D021122:Protein Subunits / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D047929:Term Birth / Q000378:metabolism; D014590:Uterine Contraction / Q000187:drug effects / Q000378:metabolism*
C121855:CACNA1G protein, human; C479952:CACNA1H protein, human; C469757:CACNA1I protein, human; D020747:Calcium Channels, T-Type; D026901:Membrane Transport Proteins; D021122:Protein Subunits; D012333:RNA, Messenger; C022838:nickel chloride; D009532:Nickel