192IgG‐saporin immunotoxin‐induced loss of cholinergic cells differentially activates microglia in rat basal forebrain nuclei

Abstract

To characterize the specificity of a novel cholinergic immunotoxin (conjugate of the monoclonal antibody 192IgG against the low-affinity nerve growth factor receptor with the cytotoxic protein saporin), coronal sections through the basal forebrain of adult rats, that received a single intracerebro-ventricular injection of 4 micrograms of 192IgG-saporin conjugate, were subjected to histochemical and immunocytochemical procedures to evaluate cholinergic (choline acetyltransferase (ChAT)-immunoreactive, acetylcholinesterase-positive, NADPH-diaphorase-positive) and GABAergic structures (parvalbumin-immunoreactive, labeling of perineuronal nets with Wisteria floribunda agglutinin) as well as microglia (visualized with Griffonia simplicifolia agglutinin) and astrocytes (immunostaining for glial fibrillary acidic protein). Seven days following injection of the immunotoxin, ChAT-immunoreactive cells nearly completely disappeared throughout the magnocellular basal forebrain complex, including globus pallidus, as compared to vehicle-injected controls. However, there was no significant difference in the number of ChAT-positive cells in the adjacent ventral pallidum and in the caudate-putamen of immunolesioned and control animals. NADPH-diaphorase-containing cells, including a significant subpopulation of cholinergic cells, also strikingly decreased in number by more than 90% in the magnocellular basal forebrain complex following immunolesion, and only a few noncholinergic diaphorase-positive cells survived in the medial septum, vertical and horizontal diagonal band, and nucleus basalis of Meynert. In contrast, the number of parvalbumin-containing GABAergic projection neurons in the septum-diagonal band of Broca complex and nucleus basalis of Meynert from immunolesioned rats was not different from that of vehicle-injected control animals. Immunolesioning also did not result in any change in either number or shape of cells surrounded by perineuronal nets, which are frequently associated with parvalbumin-containing GABAergic neurons. Seven days following injection of the immunotoxin, a very strong activation of microglia with an identical distribution pattern was observed in all experimental animals. Large numbers of activated microglia were found in all magnocellular basal forebrain nuclei, corresponding to the distribution of degenerating cholinergic cells. Additionally, immunolesioning also resulted in a dramatic activation of microglia in the lateral septal nuclei, which are known to be almost free of cholinergic cells, but not of penetrating cholinergic dendrites in adjacent zones, and in the ventral pallidum, where there was no observed loss of cholinergic cells. There was no significant increase in microglia activation in striatum and cortical areas, and no astrocytic response in any of the basal forebrain nuclei at this particular time point of survival. These results suggest that 192IgG-saporin specifically destroys basal forebrain cholinergic neurons and does not suppress their neuronal activity.(ABSTRACT TRUNCATED A...

Changes in electrocortical power and coherence in response to the selective cholinergic immunotoxin 192 IgG-saporin

“…That such cells lie outside of the septohippocampal system is suggested by our findings (Waite et al 1995), as well as those of others (Rossner et al 1995a;Zhang et al 1996) which show that 192 IgG-saporin administered i.c.v. at doses of 3.4-4.0 μg results in a near complete loss of cholinergic function in the medial septum and hippocampus as measured by ChAT assay, ChAT immunoreactivity, acetylcholinesterase, or p75 staining.…”

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.…”

“…It is worth stressing that anti-p75 NTR IgG offers a better choice for targeting viral vectors to BFCNs than NGF, because, firstly, biotinylation of NGF is known to reduce its binding activity to neurotrophin receptors [45], and, secondly, in addition to p75 NTR enriched in cholinergic neurons, NGF also binds the TrkA receptor expressed on cholinergic neurons and other cells [46,47]. 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].…”

“…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.…”