Intravenous injection of the cholinesterase inhibitor physostigimine evoked a hypertensive response in the unanesthetized 12- to 14-week-old spontaneously hypertensive rat (SHR). These responses were greatly enhanced in magnitude when compared to similar injections in normotensive Wistar-Kyoto (WKY) control rats. Stimulation of autonomic ganglia with dimethylphenylpiperazinium (DMPP) also evoked a pressor response in SHR; however, the magnitude of these responses was not different from those obtained in WKY. In unanesthetized, freely moving SHR, the intracerebroventricular (icv) injection of hemicholinium-3 (HC-3) to block the synthesis of brain acetylcholine (ACh) produced a marked hypotensive response. No reduction in arterial pressure was observed following similar injections of HC-3 in WKY. The decline in arterial pressure following icv HC-3 was correlated with the initial decline in brain ACh levels, particularly in the hypothalamus. These results indicate that enhanced sympathetic nerve activity and the resultant hypertensive state present in young SHR is dependent on increased central cholinergic activity.