Nandigama_2013_Neurosci_229_27

Nicotinic acetylcholine receptors (nAChR) influence bladder afferent activity and reflex sensitivity, and have been suggested as potential targets for treating detrusor overactivity. Mechanisms may include indirect effects, e.g. involving the urothelium, and direct action on nAChR expressed by afferent neurons. Here we determined the nAChR repertoire of bladder afferent neurons by retrograde neuronal tracing and laser-assisted microdissection/reverse transcriptase polymerase chain reaction (RT-PCR), and quantified retrogradely labelled nAChRalpha3-subunit-expressing neurons by immunohistochemistry in nAChR alpha3beta4alpha5 cluster enhanced green fluorescent protein (eGFP) reporter mice. Bladder afferents distinctly expressed mRNAs encoding for nAChR-subunits alpha3, alpha6, alpha7, beta2-4, and weakly alpha4. Based upon known combinatorial patterns of subunits, this predicts the expression of at least three basically different subunits of nAChR - alpha3( *), alpha6( *) and alpha7( *) - and of additional combinations with beta-subunits and alpha5. Bladder afferents were of all sizes, and their majority (69%; n=1367) were eGFP-nAChRalpha3 positive. Immunofluorescence revealed immunoreactivities to neurofilament 68 (NF68), transient receptor potential cation channel vanilloid 1 (TRPV1), substance P (SP) and calcitonin gene-related peptide (CGRP) in eGFP-nAChRalpha3-positive and -negative neurons. For each antigen, all possible combinations of colocalisation with eGFP-nAChRalpha3 were observed, with eGFP-nAChRalpha3-positive bladder neurons without additional immunoreactivity being most numerous, followed by triple-labelled neurons. In conclusion, more than one population of bladder afferent neurons expresses nAChR, indicating that peripheral nicotinic initiation and modulation of bladder reflexes might result, in addition to indirect effects, from the direct activation of sensory terminals. The expression of multiple nAChR subunits offers the potential of selectively addressing functional aspects and/or sensory neuron subpopulations.