The memory impairment associated with Alzheimers disease has been associated with a loss in acetylcholine function and pathophysiology of the entorhinal/perirhinal, or rhinal cortex. Infusions of a selective cholinergic immunotoxin, lead to cholinergic deafferentation of the rhinal cortex and yielded recognition deficits of the same magnitude as those produced by excitotoxic lesions of this region. Administration of the non-selective muscarinic antagonist scopolamine into the rhinal cortex results in a severe memory deficit. Scopolamine, however, includes antagonists of both m1 and m2 receptor subtypes. Therefore we compared the effects of selective infusion of the m1-selective antagonist pirenzepine, and the m2-selective antagonist methoctramine directly into the rhinal cortex. Our findings suggest that m1 and m2 receptor subtypes in the perirhinal cortex have functionally dissociable roles, and that visual recognition memory is critically dependent on the m1 receptor subtype. While the importance of the m1 receptor has been established we know very little about the m1 muscarinic-dependent intracellular signaling profiles underlying critical synaptic changes. Therefore we recently undertook a proteomics approach to uncover the molecular signaling pathways activated during memory formation in a region-specific manner using two techniques: laser capture microdissection and reverse phase protein microarrays (RPPA). Sections of snap frozen perirhinal tissue from Rhesus monkeys were prepared and stained for Nissl substance. Tissue subregions of perirhinal cortex (Layers III and V/VI), as well as hippocampus (cell body layers of CA1, CA3, and dentate gyrus and their corresponding dendritic fields), were isolated using laser capture microdissection. Lysed tissue samples were then printed onto RPPA. These arrays were probed with antibodies against phosphorylated, as well as total, proteins involved in muscarinic signaling, synaptic transmission, and neuronal activity. We are now in the process of applying this methodology to elucidate which critical proteins have been activated in specific laminae of the perirhinal cortex during visual memory formation. While the cognitive memory system of the medial temporal lobe may rely on the neuromodulator acetylcholine habit formation of the ventral neo-striatum is dependent on the neuromodulator dopamine. We recently trained monkeys on a short-ITI form of concurrent visual discrimination learning, one in which a set of stimulus pairs is repeated not only across daily sessions but also several times within each session. Learning on this faster discrimination task and not the slower learning version of the task ( a habit task) with long-ITI was impaired by systemic injections of scopolamine. The results suggest that learning discriminations with the short ITIs within a session could be driven by both cognitive memory as well habit formation working in concert. It is clear that cholinergic function plays a major role in cortical cognitive function. Groups of large cholinergic and GABAergic neurons in the BF are known to project widely throughout the cerebral cortical mantle. This input is thought to modulate cortical excitability locally, thus influencing brain functions as diverse as stimulus processing, motivation, and learning. We measured changes to spontaneous fMRI fluctuations after reversibly inactivating components of the basal forebrain, including the nucleus basalis of Meynert. Unilateral inactivation of the basal forebrain had a strong impact on spontaneous activity in the injected hemisphere. In control sessions for which saline was injected, spatial independent component analysis revealed functional networks, which were in all cases bilateral and corresponded well to the known resting state networks in the macaque. However, following inactivation, many networks became independent in the left and right cerebral cortex, indicating a hemispheric uncoupling. Furthermore, a comparison of the fMRI time series correlations across all pairs of voxels within and between hemispheres revealed that the injected hemisphere was significantly weaker in its FC than the uninjected side. These data demonstrate that resting state functional connectivity is, at least in part, shaped by long-range, common input projections arising from the basal forebrain. In a second series of experiments we examined the nature of spontaneous brain activity at rest using both fMRI and electrophysiology measurements with reversible inactivation. Initially we measured neural activity at several cortical sites while resting monkeys were scanned. The fMRI fluctuations, the neural fluctuations, and their temporal coupling, varied over time and were particularly influenced by the animals apparent state of wakefulness. In a follow-up set of experiments, we unilaterally and reversibly inactivated the Nucleus Basalis of Meynert in the basal forebrain. Following inactivation, the level of spontaneous fMRI correlation in the injected hemisphere was consistently and significantly lower than in the uninjected hemisphere. Together, the results suggest that, in the resting brain, the shared spontaneous fMRI signals that serve as the basis of functional connectivity, are closely linked in neural fluctuations, are shaped by input from the basal forebrain, and are strongly influenced by spontaneous and induced changes in behavioral arousal. Our results suggest that in addition to the important role of acytylcholine for recognition memory in the medial temporal lobe it plays a larger role in regulation of cortical activity.
