The cholinergic septo-hippocampal pathway is critically involved in learning and memory functions of the hippocampus and a likely substrate for the neurotoxic effects of low-level lead (Pb) exposure in children. We observed that exposure of neonatal rats to low levels of lead causes selective reduction in ontogenic expression of cholinergic marker enzyme cholineacetyltransferase (ChAT) and muscarinic receptors in the septum without inducing similar changes in the hippocampus. This implicates the cholinergic septo-hippocampal neurons as a particularly vulnerable target for the developmental insult by inorganic lead. In this proposal we will investigate in detail, the mechanism and functional consequences of lead's effects on the septa) cholinergic cells. (1) Receptor radioligand binding to septal homogenates from control and lead-exposed neonatal rats will be used to determine the developmental time course of the Pb-induced reduction in muscarinic receptors and its persistence into adulthood; (2) Receptor subtype-selective radioligand binding and a combination of receptor autoradiography and ChAT immunocytochemistry in brain sections and/or dissociated septal cultures will be used to determine which muscarinic receptor subtype(s) are reduced and establish their localization to the cholinergic neurons; (3) a hypothesis that Pb interferes with the regulation of receptor expression by nerve growth factor (NGF) will be tested by examining the interaction between NGF and Pb on the expression of muscarinic receptors in dissociated septal cultures: (a) 125I-NGF binding to dissociated cells will be used to determine if lead interferes with binding and uptake of NGF by the target neurons and (b) receptor hybridization will be used to determine if NGF induces and Pb inhibits the induction of receptor subtypes-specific mRNA; (4) Whole cell patch-clamp recording from dissociated septal neurons in culture will be used to determine the effect of Pb exposure on the development of chemosensitivity to ACh in the septal cholinergic neurons and test Pb's effect on the electrophysiological characteristics of these neurons; specifically, we will test a hypothesis that Pb modifies the characteristic rhythmic, pacer-like firing properties of cholinergic cells by (a) attenuating muscarinic abbreviation of postspike afterhyperpolarization (AHP) , (b) altering Ca conductances, and (c) modifying the outward Ca-dependent potassium currents in the Pb-exposed cells. The phenotype of the cells will be identified after recordings by immunocytochemical staining with anti-ChAT antibodies. These experiments should provide new insights into the neurobiology of the cholinergic septo-hippocampal neurons and the extent to which alterations in their chemosensitive and electrical membrane properties might be involved in Pb-induced behavioral toxicity.
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