We examined the effect of nitric oxide synthase (NOS) inhibition and tetrodotoxin (TTX) on the increase of cerebral blood flow (CBF) in parietal (CoBF) and cerebellar cortex (CeBF) in response to hypercapnia. Rats were anesthetized with halothane and artificially ventilated. Hypercapnia was induced by adding 5% CO2 to the inhalation mixture. CoBF and CeBF were measured by laser-Doppler flowmetry. NOS inhibition was achieved by intravenous (30 mg/kg) and/or topical application (1 mM) of NG-nitro-L-arginine (L-NNA). Activity in perivascular nerves around pial and cortical vessels was inhibited by topical application of TTX (20 microM). Under control conditions, hypercapnia (66 +/- 1 mmHg) increased CoBF by 70 +/- 4% and CeBF by 96 +/- 5%. Systemic L-NNA decreased the baseline level of CoBF and CeBF by 11 +/- 3%, but topical L-NNA did not affect baseline flow. Intravenous L-NNA attenuated the hypercapnic increase of CoBF by 77 +/- 5% and CeBF by 63 +/- 4% within 10-20 min. Topical L-NNA attenuated the hypercapnic increase of CoBF by 52 +/- 6% and CeBF by 29 +/- 5% after 45-min exposure. Both CoBF and CeBF decreased rapidly when L-NNA was infused during sustained hypercapnia, but not when L-NNA was applied topically. Effect of intravenous L-NNA was partially prevented by pretreatment with intravenous L-arginine. Intravenous or topical L-NNA enhanced the rise of CBF elicited by cortical spreading depression, adenosine (1 mM), or sodium nitroprusside (300 microM), except in the cerebellum where topical L-NNA attenuated the rise of CBF elicited by adenosine by 53%.(ABSTRACT TRUNCATED AT 250 WORDS)