The behavioral effects of two amnestic treatments (intraseptal chlordiazepoxide (CDP) and intraventricular AF64A) were examined in a delayed-nonmatch-to-sample radial-arm maze (DNMTS) paradigm. The types of errors induced by these treatments in this working memory task were assessed to determine how acute and chronic disruptions of the medial septum affect different phases of working memory (encoding, maintenance, retrieval). Rats were initially trained to perform the DNMTS task with a 1-h delay imposed between the training and the testing sessions. The first experiment demonstrated that intraseptal injection of 30 nmoles of CDP did not produce state-dependent learning. Rats were injected immediately following training with CDP or artificial cerobrospinal fluid (CSF; drug vehicle) and then prior to testing with CDP or CSF. Injection of CDP immediately following training (CDP-CSF) impaired performance of the task regardless of whether CDP was also administered before the postdelay test (CDP-CDP). Rats infused with CDP only before the postdelay test (CSF-CDP) exhibited a proactive deficit characterized by intact retention of the predelay information (i.e., arms entered prior to the delay) but impaired performance on the postdelay component (arms entered only after the delay). These data indicate: (i) that state dependency does not explain the working memory deficits induced by intraseptal CDP; (ii) that pretest CDP disrupts the storage and utilization of working memory for current arm selections. The second experiment examined the behavioral effects induced by a permanent disruption of the cholinergic septohippocampal pathway produced by icv injection of the cholinotoxin AF64A. Rats were initially trained on the DNMTS task and then bilaterally injected icv with either AF64A (2.5 nmoles/side) or CSF. AF64A-treated rats exhibited a significant impairment of performance compared to CSF-treated controls. In contrast to the impairment exhibited by CDP-treated rats in Experiment 1, the performance of AF64A-treated rats displayed a deficit in the maintenance/retrieval of information acquired during RAM training and an impairment in ability to store current spatial information in working memory to guide postdelay testing performance. These studies demonstrate that acute and chronic disruptions of the septohippocampal pathway produce distinct profiles of cognitive impairment that should help to reveal the behavioral and neurobiological characteristics of spatial memory.