Proceedings of The Physiological Society

Physiology 2015 (Cardiff, UK) (2015) Proc Physiol Soc 34, PC140

Poster Communications

Evidence of a role for TRPV2 channels in retinal arteriolar myogenic signalling

M. K. McGahon1, J. A. Fernandez1, J. D. McKee1, D. P. Dash1, A. V. Zholos2, G. J. McGeown1, T. M. Curtis1

1. Centre For Experimental Medicine, Queen's University Belfast, Belfast, select a state, United Kingdom. 2. Department of Biophysics, National Taras Shevchenko University of Kyiv, Kiev, Ukraine.

The TRP channels, TRPC6 and TRPM4, are known to play a key role in the myogenic response of cerebral arteries1. Presently, little is known about the contribution of TRP channels in mediating such responses at the level of the microcirculation. Here we show that TRPV2 is the primary TRP channel that contributes to myogenic signalling in rat retinal arterioles. Sprague-Dawley rats were killed by schedule 1 methods and retinal arterioles isolated for RT-PCR, Fura-2 Ca2+ microfluorimetry, patch-clamp electrophysiology and pressure myography studies as previously described2. In some experiments, immunohistochemistry was performed on retinal wholemount preparations2. Data were tested for normality using the D'Agostino-Pearson normality test and analysed using paired t-tests. mRNA transcripts for the mechanosensitive TRP channels, TRPC1, M7, P2 (PKD2), V1, V2 and V4, but not TRPC6 or M4, were detected in isolated retinal arterioles. Immunolabelling studies revealed cytosolic and membrane expression of TRPC1, M7, V1, V2 and P2 in retinal arteriolar myocytes, while TRPV4 appeared largely restricted to the nuclei of these cells. Hypo-osmotic stretch-induced Ca2+ influx in retinal arteriolar myocytes was reversed by the TRPV2 inhibitor tranilast (TNL; 100 μM; n=5; 100±33% vs -12.1±10.2%, P<0.05) and the non-selective TRPP2/V2 antagonist amiloride (100μM; n=8; 100±17.7% vs 63.4±12.2%, P<0.01). Inhibitors of TRPC1, M7, V1 and V4 had no effect. Hypo-osmotic stretch activated whole‑cell currents were similar in Na+ and Cs+ containing extracellular solutions (‑33.6±8.7pA/pF vs -34.0±9.8 pA/pF respectively at -80mV; P>0.05) suggesting no contribution by TRPP2 channels. Application of 35.46±2.16 mmHg of negative pressure in 14 cell-attached patches resulted in an increase in integrated current from 0.38±0.10pC/s to 2.10±4.30pC/s (P<0.001). Pressure-induced stretch failed to significantly activate current in the presence of TNL (0.42±0.24pC/s to 0.53±0.23pC/s; n=10, P>0.05). Application of TNL inhibited delta-9-tetrahydrocannabinol (10μM) activated TRPV2 currents (‑4.0±1.0pA/pF vs -0.03±0.5pA/pF at -80mV; n=6, P<0.01), but had no effect on L-Type Ca2+ channels (KCl-induced Ca2+ entry; 61.7±19.3nM vs 59.7±18.6nM in the absence and presence of TNL, respectively; n= 6, P>0.05) or ryanodine-sensitive store release (caffeine-induced Ca2+ release; 183.6±26.7nM vs 172.2±28.7nM; n=7, P>0.05). Addition of TNL to isolated pressurised (40mmHg) retinal arterioles under conditions of myogenic tone resulted in significant dilation (98.5±0.5% vs 101.9±0.7% of initial diameter upon pressurisation; n=9, P<0.001). Our results suggest that rat retinal arteriolar myocytes express a range of mechanosensitive TRP channels, but only TRPV2 appears to contribute to myogenic signalling in this vascular bed.

Where applicable, experiments conform with Society ethical requirements