Selective Immunolesions of Cholinergic Neurons in Mice: Effects on Neuroanatomy, Neurochemistry, and Behavior

Abstract

The ability to selectively lesion mouse basal forebrain cholinergic neurons would permit experimental examination of interactions between cholinergic functional loss and genetic factors associated with neurodegenerative disease. We developed a selective toxin for mouse basal forebrain cholinergic neurons by conjugating saporin (SAP), a ribosome-inactivating protein, to a rat monoclonal antibody against the mouse p75 nerve growth factor (NGF) receptor (anti-murine-p75). The toxin proved effective and selective in vitro and in vivo. Intracerebroventricular injections of anti-murine-p75-SAP produced a dose-dependent loss of choline acetyltransferase (ChAT) activity in the hippocampus and neocortex without affecting glutamic acid decarboxylase (GAD) activity. Hippocampal ChAT depletions induced by the immunotoxin were consistently greater than neocortical depletions. Immunohistochemical analysis revealed a dose-dependent loss of cholinergic neurons in the medial septum (MS) but no marked loss of cholinergic neurons in the nucleus basalis magnocellularis after intracerebroventricular injection of the toxin. No loss of noncholinergic neurons in the MS was apparent, nor could we detect loss of noncholinergic cerebellar Purkinje cells, which also express p75. Behavioral analysis suggested a spatial learning deficit in anti-murine-p75-SAP-lesioned mice, based on a correlation between a loss of hippocampal ChAT activity and impairment in Morris water maze performance. Our results indicate that we have developed a specific cholinergic immunotoxin for mice. They also suggest possible functional differences in the mouse and rat cholinergic systems, which may be of particular significance in attempts to develop animal models of human diseases, such as Alzheimer's disease, which are associated with impaired cholinergic function.

Up-Regulation of Cation-Independent Mannose 6-Phosphate Receptor and Endosomal-Lysosomal Markers in Surviving Neurons after 192-IgG-Saporin Administrations into the Adult Rat Brain

“…Additionally, a few p 75NTR -negative basal forebrain cholinergic neurons located in the ventral pallidum and ventral to the lenticular nucleus were found to be spared by the immunotoxin. This is consistent with earlier findings in rat 31 and mouse 44 and is believed to account for intact cholinergic innervations in the amygdala after 192-IgG-saporin administration.…”

Nerve Growth Factor Rapidly Increases Muscarinic Tone in Mouse Medial Septum/Diagonal Band of Broca

“…Cholinergic MS/DB neurons uniquely express p75 NTR (Berger-Sweeney et al, 2001). Although p75 NTR displays a significantly lower affinity to NGF than do Trk receptors in general, a number of roles have been ascribed to it either acting alone or in synergy with TrkA.…”

Low-Affinity Neurotrophin Receptor p75 Promotes the Transduction of Targeted Lentiviral Vectors to Cholinergic Neurons of Rat Basal Forebrain

“…Because p75 NTR was implicated in the uptake and retrograde transport of tetanus toxin and its H C fragment in motor neurons [63,64], we reasoned that the same mechanism could also be useful for vector targeting the BFCNs. After all, p75 NTR is strongly enriched in BFCNs in the adult brain, and targeting of cholinergic cells by small peptides or fluorescence probes linked to IgG 192 has been utilized for their labeling and destruction in vivo [22,27,29,30,32,65]. Our data show that conjugation of vectors with antip75 NTR antibody does, indeed, enable targeted expression of GFP-encoding vectors in BFCNs in adult rat brain.…”

“…Conjugates of anti-p75 NTR IgG with small fluorescence probes and neurotoxic peptides have been shown to target and facilitate their effects in BFCNs [29,30,48]. Hartig et al [37] also showed that Cy3 bound to IgG 192 selectively labeled BFCNs and stayed within these neurons over several months [37], while IgG 192 conjugates of ribosome inhibitor peptide saporin have been widely used for their lesions [22,27,29,30]. Importantly, neither saporin nor Cy3 linked to IgG 192 hindered its binding to p75 NTR or internalization by BFCNs.…”

“…While the functional significance of the latter has remained unclear over the years, this unique BFCN characteristic has been successfully exploited for their targeted lesion with toxins and in vivo prelabeling with fluorescence markers for behavioral and electrophysiological experiments [23][24][25][26][27][28][29][30]. To the best of our knowledge, the utility of the p75 NTR for targeting and delivery of drug candidates or vectors to BFCNs has remained unexplored.…”