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Morphological and neurochemical characterisation of anterogradely labelled spinal sensory and autonomic nerve endings in the mouse bladder

  • Harman Sharma
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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  • Melinda Kyloh
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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  • Simon J.H. Brookes
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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  • Marcello Costa
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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  • Nick J. Spencer
    Correspondence
    Corresponding authors at: College of Medicine and Public Health, Flinders University, GPO Box 2100, Adelaide, SA 5001, Australia.
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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  • Vladimir P. Zagorodnyuk
    Correspondence
    Corresponding authors at: College of Medicine and Public Health, Flinders University, GPO Box 2100, Adelaide, SA 5001, Australia.
    Affiliations
    Discipline of Human Physiology, Flinders Health and Medical Research Institute, College of Medicine and Public Health, Flinders University, Adelaide, Australia
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      Highlights

      • Immunohistochemistry was combined with anterograde labelling to identify extrinsic nerve endings in the bladder.
      • This approach distinguished spinal afferent and autonomic efferent (motor) nerve endings in the bladder.
      • Morphology alone could not reliably distinguish simple and branching endings of afferent and autonomic neurons.

      Abstract

      The bladder is innervated by axons of sympathetic and parasympathetic efferent nerves, and by spinal afferent neurons. The objective was to characterise anatomically and immunohistochemically the terminal endings of sensory and autonomic motor nerve endings in wholemount preparations of the mouse bladder. We used both anterograde labelling of pelvic and hypogastric nerves ex vivo and anterograde labelling from lumbosacral dorsal root ganglia (DRG) in vivo in male and female mice. These were combined with immunohistochemistry for major markers of sensory, sympathetic and parasympathetic nerves. Selective labelling of spinal afferent endings following dextran biotin-labelling from DRGs in vivo showed no co-localisation of VAChT or TH in sensory terminals in the detrusor and suburothelial plexus. Biotinamide was applied ex vivo to nerve trunks arising in the pelvic ganglion and running towards the bladder. Among the filled axons, 38% of detrusor fibres and 47% of suburothelial axons were immunoreactive for calcitonin-gene related peptide (CGRP). Vesicular acetylcholine transporter (VAChT) immunoreactivity was present in 26% of both detrusor and suburothelial axons. For tyrosine hydroxylase (TH), the proportions were 15% and 17%, respectively. Three major morphological types of CGRP-immunoreactive nerve endings were distinguished in the bladder wall: simple, branching and complex. VAChT-immunoreactive parasympathetic axons had simple and branching endings; TH immunoreactive axons all had simple morphologies. Our findings revealed that different subtypes of sensory and autonomic nerve endings can be reliably identified by combining anterograde labelling ex vivo with specific immunohistochemical markers, although morphologically some of these types of endings were indistinguishable.

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