Spatial distribution and developmental appearance of postjunctional P2X₁ receptors on smooth muscle cells of the mouse vas deferens

Abstract

P2X1-type purinoceptors have been shown to mediate fast transmission between sympathetic varicosities and smooth muscle cells in the mouse vas deferens but the spatial organization of these receptors on the smooth muscle cells remains inconclusive. Voltage clamp techniques were used to estimate the amplitudes of spontaneous excitatory junction currents (SEJCs) in cells of the vas deferens longitudinal smooth muscle layer. These currents involved the activation of about 6% of the P2X-type channels present on the cell, as compared to whole cell currents produced when isolated smooth muscle cells were exposed to maximal concentrations of either ATP or alpha,beta-MeATP. Immunofluorescence staining of the vas deferens with antibodies against P2X1 receptor showed a diffuse, grainy distribution over the entire membrane of each smooth muscle cell. Anti-P2X1 staining was not markedly clustered beneath anti-SV2-stained sympathetic varicosities. Similar results were obtained for cells in the urinary bladder. During development, P2X1 mRNA was detected as early as embryonic day 15 (E15). Increasing intensities of diffuse immunostaining for P2X1 were observed in the walls of the bladder, tail artery, and aorta from E15 until 6 weeks postnatal. The vas deferens showed increasing intensities of diffuse staining of its smooth muscle layers between 2 and 6 weeks postnatal, consistent with the time-course of development of fast purinergic transmission described previously. Together, the results suggest that the response of smooth muscle of the vas deferens to ATP released from sympathetic varicosities relies on rapidly desensitizing P2X1 receptors, distributed diffusely across the smooth muscle cell surface.

Neocuproine, a Copper (I) Chelator, Potentiates Purinergic Component of Vas Deferens Contractions Elicited by Electrical Field Stimulation

“…In a recent study, it was shown that ATP released by sympathetic varicosities of the mouse vas deferens binds to P2X receptors which activate fast, ligand-gated channels, resulting in depolarization of smooth muscle cells [17] . P2X1-type purinoceptors have been shown to mediate fast transmission between sympathetic varicosities and smooth muscle cells in the mouse vas deferens but the spatial organization of these receptors on the smooth muscle cells remains inconclusive [18] .…”

Correlation of non-uniform protein expression with variation in transmitter release probability

“…While variations in the post-synaptic specializations have been shown to affect synaptic strength in other terminals (e.g., Casanovas et al, 1999;Levenes et al, 2001;Lim et al, 1999), in the vas deferens only a small proportion of the P2X 1 -purinocpetors, which are evenly distributed over the entire surface of the smooth muscle (Knight et al, 2003;Lee et al, 2000;Liang et al, 2001;Vulchanova et al, 1996), are responsible for generating the spontaneous excitatory junction currents (Liang et al, 2001). This suggests that the variation in the strength of synaptic transmission in this preparation is pre-junctional in origin.…”

Increased ratio of rapsyn to ACh receptor stabilizes postsynaptic receptors at the mouse neuromuscular synapse

“…QT-6 fibroblasts were transfected as previously described (Phillips et al 1997). Whole-cell lysates were immunoblotted with antirapsyn monoclonal antibody (mAb)1234 (Froehner, 1984) using ECL-Plus chemiluminescence as described (Amersham, UK; Liang et al 2001). Expression plasmid encoding α-2,6-sialyltransferase tagged with a VSV-G epitope (Rabouille et al 1995) was a gift from Dr Rob Parton (Institute for Molecular Bioscience, St Lucia, QLD, Australia).…”