Pregnenolone sulphate- and cholesterol-regulated TRPM3 channels coupled to vascular smooth muscle secretion and contraction.
BACKGROUND
Transient receptor potential melastatin (TRPM)3 is a calcium-permeable ion channel activated by the neurosteroid pregnenolone sulfate and positively coupled to insulin secretion in beta cells. Although vascular TRPM3 mRNA has been reported, there is no knowledge of TRPM3 protein or its regulation and function in the cardiovascular system.


OBJECTIVE
To determine the relevance and regulation of TRPM3 in vascular biology.


RESULTS
TRPM3 expression was detected at mRNA and protein levels in contractile and proliferating vascular smooth muscle cells. Calcium entry evoked by pregnenolone sulfate or sphingosine was suppressed by TRPM3 blocking antibody or knock-down of TRPM3 by RNA interference. Low-level constitutive TRPM3 activity was also detected. In proliferating cells, channel activity was coupled negatively to interleukin-6 secretion via a calcium-dependent mechanism. In freshly isolated aorta, TRPM3 positively modulated contractile responses independently of L-type calcium channels. Concentrations of pregnenolone sulfate required to evoke responses were higher than the known plasma concentrations of the steroids, leading to a screen for other stimulators. beta-Cyclodextrin was one of few stimulators of TRPM3, revealing the channels to be partially suppressed by endogenous cholesterol, the precursor of pregnenolone. Elevation of cholesterol further suppressed channel activity and loading with cholesterol to generate foam cells precluded observation of TRPM3 activity.


CONCLUSIONS
The data suggest functional relevance of TRPM3 in contractile and proliferating phenotypes of vascular smooth muscle cells, significance of constitutive channel activity, regulation by cholesterol, and potential value of pregnenolone sulfate in therapeutic vascular modulation.
Naylor
Jacqueline
J
Multidisciplinary Cardiovascular Research Centre, University of Leeds, Leeds, LS2 9JT, United Kingdom.
Li
Jing
J
Milligan
Carol J
CJ
Zeng
Fanning
F
Sukumar
Piruthivi
P
Hou
Bing
B
Sedo
Alicia
A
Yuldasheva
Nadira
N
Majeed
Yasser
Y
Beri
Dhananjay
D
Jiang
Shan
S
Seymour
Victoria A L
VA
McKeown
Lynn
L
Kumar
Bhaskar
B
Harteneck
Christian
C
O'Regan
David
D
Wheatcroft
Stephen B
SB
Kearney
Mark T
MT
Jones
Clare
C
Porter
Karen E
KE
Beech
David J
DJ
D000818:Animals; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002784:Cholesterol / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects* / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D011284:Pregnenolone / Q000494:pharmacology*; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship; D050053:TRPM Cation Channels / Q000235:genetics / Q000378:metabolism*
D012333:RNA, Messenger; D050053:TRPM Cation Channels; C475667:TRPM3 protein, human; C533868:TRPM3 protein, mouse; C018370:pregnenolone sulfate; D011284:Pregnenolone; D002784:Cholesterol