The Effect Of Nitric Oxide Synthase Inhibition On The Upper Limit Of Cerebral Autoregulation In Anaesthetised Rats

EL Thompson, AM Coney, JM Marshall. University of Birmingham, Birmingham, UK

Cerebral autoregulation allows cerebral blood flow (CBF) to be maintained in the face of changes in arterial blood pressure (ABP): cerebral vasoconstriction occurs when ABP rises and vice versa.

We first aimed to develop a protocol to reliably assess the upper autoregulatory limit (the ABP at which no further constriction is possible and CBF increases) in rats by using phenylephrine (PE) to increase ABP: PE is reported to have a limited ability to cross the blood brain barrier and αα-adrenoreceptor density on cerebral vessels is low. Our primary aim was to investigate the role of nitric oxide (NO), from both neuronal and endothelial sources, in setting basal CBF and hence the vascular tone from which autoregulatory constriction occurs.

In a control group of 14 male Wistar rats (333 ±± 6g) anaesthetised with Alfaxan (17-20 mg.kg-1.h-1 i.v.), different doses of PE (0.1–60µg.kg-1min-1 i.v.) were infused sequentially to evoke graded increases in ABP. Right (R)CBF was recorded via a Transonic probe on the right common carotid artery after ligating the external carotid to minimise extra-cranial flow. In 3 subsequent groups, this protocol was performed after the neuronal nitric oxide synthase (nNOS) inhibitors Ѕ-Methyl-L-Thiocitrulline (SMTC, 0.56mg.kg-1, n=14), or 7-Nitroindazole (7-NI, dissolved in DMSO before dilution, 25mg.kg-1 i.p., final 1% DMSO, n=10), or the non-selective NOS inhibitor Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME, 10mg.kg-1, n=6). Cerebral vascular resistance (CVR) was computed as ABP/CBF. All drugs were diluted in saline and administered i.v unless stated. All procedures were carried out in accordance with UK legislation.

In the control group, RCBF achieved a plateau at 1.1±0.1ml.min-1 with initial graded doses of PE, while ABP and CVR continued to increase, demonstrating autoregulation. The upper limit of autoregulation, defined as the intersect of two linear regression lines fitted to the ‘plateau’ and ‘rising’ portions of mean CBF, was 167±3mmHg. SMTC, 7-NI and L-NAME significantly decreased baseline CBF by 0.3-0.5ml.min-1 * in each case (*: P<0.05, one-way ANOVA with post-hoc Bonferroni). In some animals of the SMTC and 7-NI groups, the upper autoregulatory limit was not significantly different from that of the control group; 172±2 (n=6) and 174±4mmHg (n=5) respectively, whereas in the remainder (n=8 and 5 in SMTC, 7-NI groups respectively), the upper limit was not reached, even at the maximal dose of PE infused. By contrast, in the L-NAME group, the upper limit was increased to188±3mmHg* in some animals (n=3) and was not reached in the remainder (n=3).

These findings indicate that PE can be used to assess the upper autoregulatory limit in the rat. The results obtained with nNOS inhibitors suggest that NO generated by nNOS exerts a tonic dilator influence on cerebral circulation and that in its absence there is greater scope for vasoconstriction such that the upper limit is raised, at least in some animals. The fact that L-NAME consistently raised or prevented reaching the upper limit, suggests that NO generated by eNOS may play an additional role in limiting cerebral autoregulatory vasoconstriction.