The expression and microanatomical localization of adenosine A1 and A3 receptor subtypes were investigated in rat pial and intracerebral arteries by immunoblotting, immunohistochemistry and in situ hybridization techniques. Pial artery membranes develop immune bands of approximately 79 and 52 kDa when exposed to anti-A1 and anti-A3 receptor protein antibodies respectively. Sympathectomy performed by bilateral superior cervical ganglionectomy did not change the pattern of adenosine A1 or A3 receptor immunochemistry. Sections of pial and intracerebral arteries processed for A1 or A3 receptor protein immunohistochemistry developed immune reaction in the tunica media, within arterial smooth muscle. A1 receptor immunoreactivity was more pronounced in large-sized compared to medium- and small-sized pial arteries and was stronger in small than in medium- or large-sized intracerebral arteries. A3 receptor immunoreactivity was more pronounced in smaller sized pial arteries compared to larger pial arteries, whereas no differences in the intensity of immune staining were noticeable between different sized intracerebral arteries. In situ hybridization histochemistry revealed a strong signal for A1 receptor and a moderate signal for A3 receptor in the tunica media of pial arteries, within smooth muscle. The present study indicates that rat pial and intracerebral arteries besides to the well characterized A2a and A2b receptors, express also A1 and A3 receptor subtypes. The identification of cerebrovascular A1 and A3 adenosine receptor subtypes may stimulate further research for detailing the mechanism(s) of regulation of cerebral circulation by adenosine.