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Functional nitrergic innervation of smooth muscle structures in the mucosa of pig lower urinary tract

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Abstract

Neurally released nitric oxide (NO) functions as an inhibitory neurotransmitter of urethral but not detrusor smooth muscles while relaxing bladder vasculature and muscularis mucosae (MM). Here, the distribution of nitrergic nerves was examined in the mucosa of pig lower urinary tract using immunohistochemistry, and their vasodilatory functions were studied by measuring arteriolar diameter changes. Properties of smooth muscle cells in the lamina propria (SMC-LP) of urethra and trigone were also investigated using florescence Ca2+ imaging. In the bladder mucosa, neuronal nitric oxide synthase (nNOS)–immunoreactive nitrergic fibres projected to suburothelial arterioles and venules. Perivascular nitrergic nerves were intermingled with but distinct from tyrosine hydroxylase (TH)–immunoreactive sympathetic or calcitonin gene–related peptide (CGRP)–immunoreactive afferent nerves. MM receive a nitrergic but not sympathetic or afferent innervation. In the mucosa of urethra and trigone, nitrergic nerves were in close apposition with sympathetic or afferent nerves around suburothelial vasculature but did not project to SMC-LP. In suburothelial arterioles of bladder and urethra, N ω-nitro-L-arginine (L-NA, 100 μM), an NOS inhibitor, enhanced electrical field stimulation (EFS)–induced sympathetic vasoconstrictions, while tadalafil (10 nM), a phosphodiesterase type 5 (PDE5) inhibitor, suppressed the vasoconstrictions. SMC-LP developed asynchronous spontaneous Ca2+ transients without responding to EFS. The spontaneous Ca2+ transients were enhanced by acetylcholine (1 μM) and diminished by noradrenaline (1 μM) but not SIN-1 (10 μM), an NO donor. In the lower urinary tract mucosa, perivascular nitrergic nerves appear to counteract the sympathetic vasoconstriction to maintain the mucosal circulation. Bladder MM but not SMC-LP receive an inhibitory nitrergic innervation.

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Abbreviations

α-SMA:

α-Smooth muscle actin

ACh:

Acetylcholine

cGMP:

Cyclic guanosine monophosphate

CGRP:

Calcitonin gene–related peptide

DSM:

Detrusor smooth muscle

EFS:

Electrical field stimulation

LUTS:

Lower urinary tract symptoms

L-NA:

N ω-nitro-L-arginine

MM:

Muscularis mucosae

nNOS:

Neuronal nitric oxide synthase

NO:

Nitric oxide

PDE5:

Phosphodiesterase type 5

PSS:

Physiological salt solution

SMC:

Smooth muscle cell

SMC-LP:

Smooth muscle cell in the lamina propria

TH:

Tyrosine hydroxylase

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Acknowledgements

The authors wish to thank Dr Richard Lang (Monash University) for critical reading of the manuscript.

Funding

The present study was partly supported by Grant-in-Aid for Scientific Research (C) from Japan Society for Promotion of the Science to R.M. (No. 19K08426) and H.H. (No. 17K11187, 20K09564).

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Authors and Affiliations

  1. Department of Cell Physiology, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan

    Retsu Mitsui, Yota Chikada, Keiji Arai & Hikaru Hashitani

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  1. Retsu Mitsui
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  2. Yota Chikada
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  3. Keiji Arai
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  4. Hikaru Hashitani
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Correspondence to Retsu Mitsui.

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Ethical approval

The experimental protocols used in the present study were approved by the animal experimentation ethics committee at Nagoya City University Graduate School of Medical Sciences (No. H-30 M-44).

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Supplementary Information

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Supplementary file1 Spontaneous Ca2+ transients in SMC-LP of the urethra corresponding to Ca2+ fluorescence images in Fig. 10a. (MP4 2561 KB)

Supplementary file2 Spontaneous Ca2+ transients in SMC-LP of the trigone corresponding to fluorescence images in Fig. 10b. (MP4 2200 KB)

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Mitsui, R., Chikada, Y., Arai, K. et al. Functional nitrergic innervation of smooth muscle structures in the mucosa of pig lower urinary tract. Cell Tissue Res 386, 513–531 (2021). https://doi.org/10.1007/s00441-021-03521-9

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