1. In rat hippocampal pyramidal cells in vitro, a brief train of action potentials elicited by direct depolarizing current pulses injected through an intracellular recording electrode is followed by a medium-duration afterhyperpolarization (mAHP) and a longer, slow AHP. We studied the mAHP with the use of current-clamp techniques in the presence of dibutyryl cyclic adenosine 3',5'-monophosphate (cAMP) to block the slow AHP and isolate the mAHP. 2. The mAHP evoked at hyperpolarized membrane potentials was complicated by a potential generated by the anomalous rectifier current, IQ. The mAHP is insensitive to chloride ions (Cl-), whereas it is sensitive to the extracellular potassium concentration ([K+]o). 3. At slightly depolarized levels, the mAHP is partially Ca2+ dependent, being enhanced by increased [Ca2+]o and BAY K 8644 and depressed by decreased [Ca2+]o, nifedipine, and Cd2+. The Ca2(+)-dependent component of the mAHP was also reduced by 100 microM tetraethylammonium (TEA) and charybdotoxin (CTX), suggesting it is mediated by the voltage- and Ca2(+)-dependent K+ current, IC. 4. Most of the Ca2(+)-independent mAHP was blocked by carbachol, implying that IM plays a major role. In a few cells, a small Ca2(+)- and carbachol-insensitive mAHP component was detectable, and this component was blocked by 10 mM TEA, suggesting it was mediated by the delayed rectifier current, IK. The K+ channel antagonist 4-aminopyridine (4-AP, 500 microM) did not reduce the mAHP. 5. We infer that the mAHP is a complex potential due either to IQ or to the combined effects of IM and IC. The contributions of each current depend on the recording conditions, with IC playing a role when the cells are activated from depolarized potentials and IM dominating at the usual resting potential. IQ is principally responsible for the mAHP recorded at hyperpolarized membrane potentials.