William F. Childres

Stimulation of α2 Adrenoceptors Dilates the Rat Middle Cerebral Artery

Background Because alpha2 adrenoceptor agonists are used as adjuncts to anesthetics, their effects on the cerebrovascular circulation are of prime importance. We studied changes in the diameter of rat middle cerebral arteries after stimulation of alpha2 adrenoceptors with UK14,304. Methods Rat middle cerebral arteries were isolated, cannulated at each end with a glass micropipette, and pressurized to 85 mmHg. The middle cerebral arteries were immersed in a bath (37 degrees C) containing physiologic saline solution, and luminally perfused with physiologic saline solution (100 micro liter/min). Changes in vessel diameter were measured after magnification with a microscope. Results Resting diameter of the middle cerebral arteries was 239 +/-13 micro meter (n = 8) for the first study. A dose-dependent dilation was produced by addition of UK14,304 to the extraluminal bath; a 10-15% increase in diameter occurred at a concentration of 10 sup -4 M. The dilations produced by UK14,304 were blocked with selective alpha2 -antagonists, idazoxan and rauwolscine, but not by the selective alpha sub 1 -antagonist, prazosin. The dilations could be blocked by removal of the endothelium, or the nitric oxide synthase inhibitor, N-nitro-L-arginine methyl ester (10 sup -5 M). The inhibitory effects of N-nitro-L-arginine methyl ester were reversed with the addition of 10 sup -3 M L-arginine, but not 10 sup -3 M D-arginine. Furthermore, the dilation produced by UK14,304 was completely abolished with pertussis toxin (100 ng/ml). Conclusions It was concluded that the stimulation of alpha2 adrenoceptors with UK14,304 produced a dilation in the rat middle cerebral artery that (1) was dependent on intact endothelium, (2) involved nitric oxide, and (3) acted via a pertussis toxin-sensitive G protein.

Inward rectifier potassium channels in the rat middle cerebral artery.

Inward rectifier K+ channels (Kirs) were studied in the isolated perfused rat middle cerebral artery (MCA). The addition of 15 mM K+ (KCl) to the extraluminal bath dilated the MCAs. These dilations were blocked by selective inhibitors for the Kirs (40 μM BaCl2 or 40 mM CsCl) but not selective inhibitors for other K+channels (glibenclamide, tetraethylammonium, or 4-aminopyridine). Neither removal of the endothelium nor treatment with the nitric oxide synthase inhibitor ( N G-nitro-l-arginine methyl ester, 10 μM) affected the K+-induced dilation. The addition of BaCl2 to resting MCAs produced a dose-dependent constriction of 8-12%, indicating that, during resting conditions, Kirs aid in setting or determining the resting tone. The magnitude of the dilations produced by the addition of K+ or constrictions produced by BaCl2 were independent of pressure over a range of 40-100 mmHg. We conclude that Kirs, which produce a dilation when activated, exist on the vascular smooth muscle of the rat MCA. These Kirs aid in determining the resting tone of the vessel, and their function is independent of pressure over physiological pressure ranges.

Altered Function of Inward Rectifier Potassium Channels in Cerebrovascular Smooth Muscle After Ischemia/Reperfusion

BACKGROUND AND PURPOSE Several recent studies have demonstrated that inward rectifier potassium channels (K(ir)s) are located on vascular smooth muscle of cerebral arteries in the rat. Activation of the K(ir)s dilates the arteries by relaxing the vascular smooth muscle. We tested the following hypothesis in the present study: function of inward rectifier potassium channels is altered after ischemia/reperfusion (I/R). METHODS Temporary (2-hour) focal ischemia was induced in male Long-Evans rats (3% isoflurane anesthesia) by the intraluminal filament model. After 24 hours of reperfusion, ipsilateral and contralateral middle cerebral arteries (MCAs) were harvested and mounted on micropipettes, pressurized to 85 mm Hg, and luminally perfused. RESULTS Resting diameters for contralateral (control) and ipsilateral (I/R) MCAs were not significantly different (215+/-4 microm and 211+/-5 microm [n = 6 and n = 7], respectively). Activation of the K(ir)s by abluminal administration of 15 mmol/L KCl to the control MCAs dilated the MCA by 34+/-4% (n = 8). Activation of the K(ir)s in I/R MCAs produced a dilation of only 11+/-3% (n = 8; P<0.001 compared with control). BaCl2 (75 micromol/L), a concentration-selective inhibitor of the K(ir)s, significantly attenuated the dilation produced by 15 mmol/L KCl in control MCAs but not in the I/R MCAs. Endothelial-mediated dilations elicited by the luminal administration of uridine triphosphate (10 micromol/L) produced similar dilations in both groups (32+/-5% for sham [n = 4] and 33+/-2% for I/R [n = 4]), indicating that dilator function in general was not altered in I/R vessels. CONCLUSIONS We conclude that Kir function is altered after I/R. The Kir altered function is likely to exacerbate the brain injury occurring after I/R.

Effect of Decreased Glucose Concentration on Cerebrovascular Tone In Vitro

We studied the effect of decreased glucose concentration on cerebrovascular tone in vitro. Segments of rat middle cerebral arteries (MCA) were isolated, cannulated at both ends with glass micropipettes, and pressurized to 85 mm Hg. Decreasing the glucose in the extraluminal bath and luminal perfusate from 5.5 mmol/L to 1.0 or 0.5 mmol/L for 1.5 hours each had no significant effect on the diameter of the arteries. When all the glucose was removed from the extraluminal bath and luminal perfusate for 1.5 hours, the MCA dilated by 23% [252 ± 24 (SD) μm to 311 ± 7 μm (P < .5, n = 7)]. This dilation was 80% of the maximum dilation produced by removal of Ca+2 from the bathing solutions. Neither removal of the endothelium nor inhibition of the ATP-sensitive K channels with 10−5 mol/L glibenclamide altered the response of the isolated MCA to the removal of glucose. We conclude that rat MCA are relatively more resistant to substrate limitation compared to the brain as a whole.

ENDOTHELIAL-MEDIATED DILATIONS FOLLOWING SEVERE CONTROLLED CORTICAL IMPACT INJURY IN THE RAT MIDDLE CEREBRAL ARTERY

The mechanisms associated with dysfunction of the cerebral vasculature following head trauma have not yet been fully elucidated. In an attempt to shed more light on the matter, we investigated the endothelial-mediated dilations in the rat middle cerebral artery (MCA) following severe traumatic brain injury (TBI). Rats were subjected to severe controlled cortical impact injury (CCI; 5 m/s, 130 ms duration, 3 mm deformation) over the right parietal cortex. At 24 h postinjury, ipsilateral segments of MCA and corresponding contralateral segments were isolated, mounted in a vessel chamber, and pressurized. The responses to 2 methylthio-ATP (2MeSATP), a selective agonist for the P2Y1 purinoceptors, N(omega)-nitro-L-arginine (L-NAME), an NO synthase inhibitor, and S-nitroso-N-acetylpenicillamine (SNAP), an exogenous NO donor, were determined. 2MeSATP elicited concentration dependent dilations in all MCAs studied. Ipsilateral MCAs harvested following TBI or sham-TBI, showed similar maximum dilations to 2MeSATP [70 +/- 4% (n = 17) and 72 +/- 6% (n = 13), respectively]. However, TBI reduced the concentration of 2MeSATP necessary to elicit one-half of the maximum dilation (EC50) from 15 to 9 nM (p < 0.05). Inhibition of NO synthase with 10(-5) M L-NAME abolished the dilation to 2MeSATP in both TBI and sham-TBI MCAs. The constriction to L-NAME was significantly reduced in TBI MCAs compared to sham vessels. Dilations to SNAP, an NO donor, were not altered by TBI indicating that the mechanisms of dilation involving NO in the vascular smooth muscle were not affected. Unlike other pathological conditions which often diminish endothelial-mediated responses, severe TBI enhanced the sensitivity to 2MeSATP without altering the maximum response.