Localization of TRPC1 channel in the sinus endothelial cells of rat spleen

Abstract

The ultrastructural localization of transient receptor potential C1 (TRPC1) channels in the sinus endothelial cells of rat spleen was examined by confocal laser scanning and electron microscopy. In addition, the localization of the closely associated proteins and channels, VE-cadherin, calreticulin, inositol-1,4,5-trisphosphate receptors type 1 (IP3R1), and ryanodine receptor (RyR), was also examined. Immunofluorescence microscopy of tissue cryosections revealed TRPC1 channels to be localized within the cytoplasm, in the superficial layer of the apical and basal parts of the cells, and in the junctional area of the adjacent endothelial cells. The distribution of Ca2+-storing tubulovesicular structures within endothelial cells was established by using tissue sections treated with osmium ferricyanide. Electron microscopy revealed densely stained tubulovesicular structures closely apposed to the plasma membrane and that occasionally ran closely parallel to the plasma membrane and near the caveolae and junctional apparatus. Immunolocalization analysis at the electron microscopy level using immunogold bound to the secondary antibody confirmed that TRPC1 channels were localized in the plasma membrane, caveolae, and vesicular structures in the subplasmalemmal cytoplasm of sinus endothelial cells. Calreticulin was predominantly localized in endoplasmic reticulum. IP3R1 and RyR, considered to be type 3, were colocalized in endoplasmic reticulum in proximity to the plasma membrane and caveolae. Thus, TRPC1 channels in sinus endothelial cells of the spleen might play an important role in controlling blood cell passage through phenomena including cytoskeletal reorganization, cell retraction, and disassembly of adherens junctions.

Transient Receptor Potential Channel 1 (TRPC1) Reduces Calcium Permeability in Heteromeric Channel Complexes

“…In our study the lack of agonist-, GTP␥S-, or phosphatidylinositol 4,5-bisphosphate depletion-induced currents in TRPC1 overexpressing cells suggests that at least in the heterologous expression system employed by us, TRPC1 proteins per se are unable to form functional homomeric channels in the plasma membrane. This might be explained by their inability for homotetramerization or by intracellular localization of TRPC1 homomers, preventing insertion into the plasma membrane (35)(36)(37). However, at present we cannot rule out that homomeric TRPC1 channels are not gated by typical TRPC activators.…”

Recent Developments in Vascular Endothelial Cell Transient Receptor Potential Channels

“…104 Other studies have identified TRPC1 and TRPC4 within the caveolae of rat spleen sinus endothelial cells and mouse interstitial cells of Cajal, respectively. 8,24 Recently, TRPC1 and TRPV6 has been reported to…”

“…In another example, TRPC1 is expressed in the caveolae of rat spleen sinus endothelial cells. 24 Caveolae may only express a limited number of TRP isoforms. 107 Therefore, activation of TRPC1 may be associated with caveolae-related functional changes, such as modulation of endothelial NO synthase and VRAC, but the activation of TRP channels that are not present in the caveolae will have different functional consequences.…”

“…TRPC1 was found to be localized in the plasma membrane, caveolae, and lateral plasma membranes of adjoining endothelial cells of sinus endothelial cells in rat spleen. 24 It is reasonable to speculate that TRPC1 in the lateral plasma membrane of the adjoining endothelial cells may participate in the disassembly of vascular endothelial cadherin at the adherens junctions and that the TRPC1 in caveolae may regulate endothelial NO synthase and VRAC. In this way, the signaling pathway downstream of a particular TRP isoform diverges to initiate multiple functional responses.…”

Transient Receptor Potential Channels and Caveolin-1: Good Friends in Tight Spaces: Fig. 1.

“…G protein-coupled receptors (GPCRs), heterotrimeric G proteins, receptor tyrosine kinases, Rho-1, calmodulin, VAMP-2 syntaxin, ezrin, the Na ϩ -H ϩ exchange regulatory factor (NHERF), Z01, Rac-1, stathmin, soluble N-ethylmaleimide-sensitive factor attachment protein receptor proteins, adenylyl cyclase, phosphatidyl inositol-3 kinase, phospholipase C (␤ and ␥), cytoskeletal protein 4, endothelial nitric-oxide synthase, and ion channels are examples of the signaling proteins and molecules known to coexist with caveolin in caveolae (Lockwich et al, 2000;Ostrom et al, 2002;Brazer et al, 2003;Beech, 2005;Cioffi et al, 2005;Ambudkar, 2006;Swaney et al, 2006). Transient receptor potential (TRP) channels have recently been reported to localize within caveolae, particularly the canonical TRPC1, TRPC3, and TRPC4 isoforms (Lockwich et al, 2001;Torihashi et al, 2002;Brazer et al, 2003;Nilius et al, 2003;Uehara, 2005). TRPC1 also coimmunoprecipitates with caveolin-1 (cav-1), one of the three caveolin isoforms (Lockwich et al, 2000;Bergdahl et al, 2003), even more suggestive that it exists within caveolae.…”