Cholinergic input to cerebral cortex, which largely originates in the basal forebrain, appears to be involved in arousal and memory and is greatly reduced in Alzheimer's disease. Although the cholinergic innervation is distributed throughout cortex, it is concentrated particularly in layer 5, the major source of cortical efferents. We have developed techniques for a) labelling specific classes of cortical neurons by retrograde filling, in particular layer 5 corticocollicular neurons, and b) dissociating postnatal cortical or basal forebrain neurons and maintaining the neurons in tissue culture under conditions where the labelled cells can be identified over several weeks. We have shown that the labelled neurons can be penetrated with micropipettes for recording pharmacological responses and the """"""""driver/follower"""""""" recordings, including a labelled cell and a nearby cell with which it is synaptically connected, can be obtained. With this approach we have also succeeded in establishing criteria for identifying inhibitory Gabaergic neurons in the cultures.
The aim of this proposal is to use our tissue culture preparation to characterize interactions of cholinergic basal forebrain neurons with identified cortical cells. In cultures prepared from postnatal rats, layer 5 corticocollicular neurons, previously labelled in vivo by retrograde transport, Gabaergic interneurons, identified by physiological criteria, and cholinergic basal forebrain neurons, labelled by retrograde transport, will be penetrated for intracellular recording with micropipettes. The pharmacology and ionic basis of the responses of these cells will be investigated with bath applied agonists and with synaptically driven postsynaptic responses. To study the pharmacology of synaptic interactions between cholinergic basal forebrain neurons and cortical neurons, co-cultures of basal forebrain and cerebral cortex will be prepared. Both cholinergic input from basal forebrain neurons onto cortical cells and excitatory amino acid input from cortical pyramidal cells onto basal forebrain neurons will be investigated. In a parallel set of experiments, the possibility that cortical cells provide NGF-like trophic support for cholinergic basal forebrain neuron will be tested in tissue culture. These experiments will increase our understanding of how cholinergic basal forebrain cells interact with cerebral cortex and thus are important for gaining a better understanding of the etiology of Alzheimer's disease.
