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Expression of Pcp2 in the cerebellum of Gnao−/− mice. (PDF 380 kb)

The gross anatomy of the cerebellum is normal in Gnao−/− mice. (PDF 187 kb)

PFs in the ML visualized in vGluT1-stained cerebellum. (PDF 85 kb)

We have previously provided pharmacological evidence that stimulation of calcium-sensing receptors (CaSR) induces endothelium-dependent relaxations of rabbit mesenteric arteries through activation of heteromeric TRPV4/TRPC1 channels and nitric oxide (NO) production. The present study further investigates the role of heteromeric TRPV4/TRPC1 channels in these CaSR-induced vascular responses by comparing responses in mesenteric arteries from wild-type (WT) and TRPC1-/- mice. In WT mice, stimulation of CaSR induced endothelium-dependent relaxations of pre-contracted tone and NO generation in endothelial cells (ECs), which were inhibited by the TRPV4 channel blocker RN1734 and the TRPC1 blocking antibody T1E3. In addition, TRPV4 and TRPC1 proteins were colocalised at, or close to, the plasma membrane of endothelial cells (ECs) from WT mice. In contrast, in TRPC1-/- mice, CaSR-mediated vasorelaxations and NO generation were greatly reduced, unaffected by T1E3, but blocked by RN1734. In addition, the TRPV4 agonist GSK1016790A (GSK) induced endothelium-dependent vasorelaxations which were blocked by RN1734 and T1E3 in WT mice, but only by RN1734 in TRPC1-/- mice. Moreover, GSK activated cation channel activity with a 6pS conductance in WT ECs but with a 52 pS conductance in TRPC1-/- ECs. These results indicate that stimulation of CaSR activates heteromeric TRPV4/TRPC1 channels and NO production in ECs, which are responsible for endothelium-dependent vasorelaxations. This study also suggests that heteromeric TRPV4-TRPC1 channels may form the predominant TRPV4-containing channels in mouse mesenteric artery ECs. Together, our data further implicates CaSR-induced pathways and heteromeric TRPV4/TRPC1 channels in the regulation of vascular tone.

The high sensitivity of scotopic vision depends on the efficient retinal processing of single photon responses generated by individual rod photoreceptors. At the first synapse in the mammalian retina, rod outputs are pooled by a rod "ON" bipolar cell, which uses a G-protein signaling cascade to enhance the fidelity of the single photon response under conditions where few rods absorb light. Here we show in mouse rod bipolar cells that both splice variants of the G(o) α subunit, Gα(o1) and Gα(o2), mediate light responses under the control of mGluR6 receptors, and their coordinated action is critical for maximizing sensitivity. We found that the light response of rod bipolar cells was primarily mediated by Gα(o1), but the loss of Gα(o2) caused a reduction in the light sensitivity. This reduced sensitivity was not attributable to the reduction in the total number of G(o) α subunits, or the altered balance of expression levels between the two splice variants. These results indicate that Gα(o1) and Gα(o2) both mediate a depolarizing light response in rod bipolar cells without occluding each other's actions, suggesting they might act independently on a common effector. Thus, Gα(o2) plays a role in improving the sensitivity of rod bipolar cells through its action with Gα(o1). The coordinated action of two splice variants of a single Gα may represent a novel mechanism for the fine control of G-protein activity.

12 páginas, 11 figuras, 2 tablas -- PAGS nros. 277-298, Supported in part by NIH Grant AM-19318 to L.B.