The Role of Endothelium and Nitric Oxide in Rat Pial Arteriolar Dilatory Responses to CO ₂ in vivo

Using a closed cranial window system and intravital microscopy/videometry, we studied the rat pial arteriolar (30–60 μm) responses to CO 2 before and following a light/dye (L/D) endothelial injury or topical application of the nitric oxide synthase (NOS) inhibitor, nitro-l-arginine (L-NA) or its inactive form, D-NA. L/D treatment consisted of intravenous injection of sodium fluorescein and the illumination (for 90 s) of arteriolar discrete segments on the cortical surface with light from a mercury lamp. Functional changes in pial arteriolar endothelium were characterized by evaluating responses to topical application of acetylcholine (Ach, 5 × 10 −4 M) and to intravenous (i.v.) oxotremorine (OXO, a stable blood–brain barrier permeant muscarinic agonist, 1 μg kg −1 min −1 ). After the L/D injury, dilation to Ach was absent whereas dilations to the NO donor, S-nitrosoacetyl-penicillamine (SNAP, 10 −5 M) and to CO 2 (5%) were unchanged (P a co 2 = 70 mm Hg). Loss of Ach response but intact SNAP response confirmed functional endothelial injury and intact smooth-muscle function. The global endothelium-dependent vasodilation induced by i.v. OXO was markedly attenuated when expanding the L/D injury field from 300 μm to 6 mm in diameter. However, the global vasodilation induced by inhalation of CO 2 was still unaffected by this increase in the area of light exposure. This provides evidence that the expanded exposure was capable of impairing global vasodilation resulting from endothelium-dependent stimuli but not from inhalation of CO 2 . The intact CO 2 response despite an endothelial dysfunction suggests that the reported NO dependence of hypercapnia-induced cerebral hyperemia in rats cannot be attributed to an endothelial NO source. Topical suffusion of L-NA (1 m M) for 45–60 min in our preparation blocked the pial arteriolar response to Ach, whereas CO 2 and SNAP responses were unaffected. An attenuation (by 50%) of the response to CO 2 was achieved if suffusion of L-NA was given for ≥2 h. Suffusion of D-NA, applied in the same manner, did not influence responses to any of the above applications. This demonstrates that there is a NO-dependent component for hypercapnic cerebral vasodilation even at the pial arteriolar level. The strikingly different time-related effect of topical L-NA on the Ach and CO 2 responses, together with the lack of effect of endothelial injury on CO 2 -induced dilation, strongly suggest a nonendothelial source of NO in hypercapnic cerebrovascular dilation.