We investigated the functional heterogeneity of cerebral pial arteries that are extrinsically innervated versus penetrating brain parenchymal arterioles (PA) that are intrinsically innervated by comparing myogenic activity and reactivity to neurotransmitter. Pial middle cerebral arteries (MCA, n = 6) and PA (n = 6) that branched off the MCA and penetrated the brain tissue were isolated from male Wistar rats and studied in vitro under pressurized conditions for reactivity to serotonin (5-hydroxytryptamine, 5-HT), noradrenaline (NA), and indolactam-V (IL-V), a protein kinase C (PKC) agonist. In a separate group of vessels from the same locations (n = 12), perivascular nerve density was determined after staining for protein gene product 9.5 (PGP 9.5). PAs were significantly smaller than MCAs, and possessed greater myogenic tone at all pressures studied. MCAs reacted to both 5-HT and NA with concentration-dependent contraction, however, PA had little to no response to either neurotransmitter. The percent constriction to 5-HT and NA for MCA versus PA at the maximum concentration was: 31 ± 6% versus 1.0 ± 1.0% and 13 ± 5% versus 2.6 ± 1.8% (P < 0.01). However, both types of vessels contracted with similar reactivity to PKC activation with IL-V (41 ± 4% versus 37 ± 7%, ns). Perivascular nerve density correlated with reactivity such that MCAs were densely innervated with varicose fibers within the adventitia; however, PA had very few or no adventitial fibers. The differential response to neurotransmitter suggests that there is significant heterogeneity in the cerebral circulation. It appears that in PA, the dominant vasoconstricting stimulus is intrinsic myogenic tone and that the role of neurotransmitter and intrinsic innervation is beyond that of controlling CBF.