Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, PC236

Poster Communications

Mechanisms of glutamatergic regulation of capillary diameter by pericytes

C. Reynell1, C. N. Hall1, D. Attwell1

1. Neuroscience, Physiology & Pharmacology, University College London, London, United Kingdom.

Neurovascular coupling mechanisms increase the blood flow to regions where neurons are active. Arterioles dilate to increase blood flow in response to neuronal activity, but blood flow may also be regulated by contractile pericytes on capillary walls, which dilate in response to glutamate in brain slices (Peppiatt et al., 2006). The aim of this study was to investigate the mechanisms underlying the dilation to glutamate in capillaries, and thus to better understand the contribution of pericytes to regulating blood flow. Capillaries were monitored in the molecular layer of juvenile (P12) rat cerebellar slices (200 μm) using bright field imaging. Slices were superfused with ACSF solution and oxygenated by bubbling with 95% O2/5% CO2, pH 7.4, 32-35oC. Pericytes were identified from their position on the outside of capillaries (defined as vessels less than 10 μm in diameter lacking a continuous layer of smooth muscle), and in some experiments by the expression of DsRed under control of the NG2 promoter. After preconstricting pericytes with 2 μM noradrenaline (which produced a decrease in vessel diameter of 14.4±1.8%; mean+s.e.m, n=131 capillaries; Wilcoxon signed-rank test, p<0.001), exogenous glutamate (500 μM) dilated capillaries by 7.1±1.6% (n=131; Wilcoxon signed-rank test, p<0.001), and this dilation was not reduced by blocking action potentials with 1 μM TTX (Mann-Whitney U-test, p=0.054). In TTX, glutamate produced a 30.6±10.9% increase in vessel diameter (n=5; paired t-test, p=0.048). A dilation of 8.7±1.2% (n=50; Wilcoxon signed-rank test, p<0.001) also occurred (with a latency of ~6 sec) when endogenous glutamate was released by stimulating the parallel fibres. The glutamate-evoked dilation was blocked by inhibiting nitric oxide (NO) release with 100 μM L-nitroarginine (Mann-Whitney U-test, p=0.04). In L-nitroarginine, glutamate produced a -2.0±4.5% change in vessel diameter (n=46; Wilcoxon signed-rank test, p=0.84). Interestingly, this block of glutamate dilation by L-nitroarginine was absent if synthesis of the vasoconstricting arachidonic acid derivative 20-HETE was also prevented using 1 μM HET-0016 (Mann-Whitney U-test, p=0.006 comparing glutamate in L-nitroarginine with and without HET-0016). In L-nitroarginine plus HET-0016 glutamate produced a 24.0±8.8% increase in vessel diameter (n=10; paired t-test, p=0.048). This implies that NO acts by suppressing 20-HETE synthesis, and the continued presence of dilation when NO and 20-HETE production are blocked suggests that glutamate-evoked dilation is produced by other mechanisms, such as activity-evoked release of prostaglandin or EET derivatives of arachidonic acid.

Where applicable, experiments conform with Society ethical requirements