Immunohistochemical localization of α1 nicotinic acetylcholine receptors in lymphoid tissues of humans

Abstract

Although neuroanatomical studies have indicated the occurrence of nicotinic acetylcholine receptors (nAChR) in mononuclear lymphocytes of humans, no specific subtypes of nAChR or their distribution in lymphoid tissue have been investigated. The identification of α1 nAChR in immune tissues of post mortem samples was done by immunohistochemistry using antibodies against the subunits of nAChRs. The tissues were processed for Hematoxillin and Eosin staining and indirect immunohistochemistry. Monoclonal anti-nAChR (α1 subunit) raised in rat was applied to label the tissues and linked to biotinylated anti-rat IgG. Labeled Streptavidin Biotin method was applied with Diaminobenzidene as chromogen. Skeletal muscle (positive to α1nAChR) was processed as control. The microscopic images of immunostained slides were computerized for digital image analysis. The intensity of the staining was determined based upon a score of 0, 1+ (focal staining, > 10% cells), 2+ (focal to diffuse staining, 10% > 50% cells), 3+ (diffuse staining, 50>100% of cells). Diffused (1+) distribution of α1nAChR was observed in the parenchyma of liver. Similar distribution of α1 nAChR was observed in Peyers patches and the lymphoid aggregation of the posterior part of the tongue. The capsule and red pulp areas of spleen were highly immunoreactive to anti-α1nAChRs, while a low grade immunoreactivity (IR) was observed in periarteriolar lymphoid aggregations and germinal centres. In lymph nodes, the sub scapsular sinus, medullary cords and trabeculae were intermediately IR to α1 nAChR, while in lymphoid follicles it was absent. Overall, the α1 nAChR IR was high in regions predominantly having T cells and macrophages, and it was low in regions having B cell subsets of lymphoid tissues. These findings confirm that the neuroimmune modulation could be brought by the presence of parasympathetic nerves in lymphoid tissues through α1nAChRs. Further investigations are needed using antibodies against different cholinergic components to confirm the complete architecture of cholinergic nerve supply in immune tissues which would be essential in understanding the neuro immune modulation.