The expression of ß3-adrenoceptor and muscarinic type 3 receptor immuno-reactivity in the major pelvic ganglion of the rat.

Bladder afferent outflow, linked to sensation, plays a critical role in bladder pathology: abnormal outflow results in altered sensation, leading to increased voiding frequency, urge and often incontinence. ß3-adrenoceptor agonists have been suggested to be beneficial in treating these symptoms. However, the absence of a significant sympathetic innervation of the detrusor and only a modest relaxation of bladder muscle by ß3 agonists has questioned the therapeutic site of action of ß3 agonists in the bladder. The present study was done to explore the possibility that ß3-adrenoceptors might be located in the pelvic plexus. Using the rat, where the pelvic plexus is located primarily within a single ganglion, the major pelvic ganglion (MPG), immuno-histochemical approaches were used to identify structures expressing ß3-adrenoceptor immuno-reactivity (ß3AR-IR). The only structures found to express ß3AR-IR were small-diameter tyrosine hydroxylase and vesicular mono-amine transporter immuno-reactive (TH-IR and vmat-IR) neurones. These neurones, found in clusters or singly on the periphery of the ganglion, or dispersed in smaller clumps throughout the MPG, are similar to the small intensely fluorescent (SIF) cells described previously. Not all small cells expressed ß3AR-IR. A population of the small cells were also immuno-reactive to the type 3 muscarinic receptor (M3R-IR) and the P2X3 purinergic receptor (P2X3-IR). Clumps of small cells were associated with calcitonin gene-related peptide immuno-reactive (CGRP-IR) nerve fibres (putative sensory fibres) and a small number were contacted by putative cholinergic nerves expressing immuno-reactivity to vesicular acetylcholine transporter (vacht-IR). These observations are consistent with the idea that small cells are interneurons and one of the components making up complex neural circuits within the MPG. The precise physiological role of these neural elements in the MPG is unknown. However, as one therapeutic action of ß3-adrenoceptor agonists is to modulate sensation, it is possible that these neural circuits may be involved in the regulation of afferent outflow and sensation.

Demonstration of intrinsic innervation of the guinea pig upper urinary tract using whole-mount preparation.

AIMS: The morphology and functional importance of the autonomic nervous system in the upper urinary tract is still not completely understood. Previous histological studies investigating the innervation of the urinary tract have mainly used conventional sections in which the three-dimensional structure of the intramural innervation is difficult to achieve. In contrast, the whole-mount preparation technique is a suitable method for visualizing the distribution of the mesh-like neuronal networks within the urinary tract. METHODS: The distribution and regional variation of neurofilament (NF), tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), and substance P-immunoreactive (SP-IR) neurons, as well as acetylcholinesterase (AChE) and nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d)-positive neurons were investigated using whole-mount preparations of the guinea pig upper urinary tract. RESULTS: Two distinct nervous plexuses were detected within the muscle layers containing NF, TH, ChAT, and SP-IR nerves. AChE-positive nerves were seen in all layers. Only moderate NADPH-d-positive innervation was found. Renal pelvis, upper and lower part of the ureter showed an overall increased innervation compared to the middle portion of the ureter. Ganglia were found at the pelviureteric border displaying NF and TH immunoreactivity. CONCLUSION: The whole-mount preparation technique provides an elegant method for assessing the three-dimensional architecture of ureteral innervation. The guinea pig upper urinary tract is richly supplied with adrenergic, cholinergic, nitrergic, and sensory nerves which suggest that the autonomous nervous system plays an important role in controlling ureteral motility and blood flow.

Localisation of NADPH-diaphorase and acetylcholinesterase activities and of tyrosine hydroxylase and neuropeptide-Y immunoreactivity in neurons of the hypogastric ganglion of young adult and aged rats.

Neurons in the hypogastric (main pelvic) ganglia of 4- and 24-month-old male rats were investigated by enzyme histochemical methods for NADPH-diaphorase (NADPH-d) and acetylcholinesterase (AChE) activities and by immunofluorescence for tyrosine hydroxylase (TH) and neuropeptide Y (NPY) immunoreactivities. Systematic random sampling of standard sized areas of ganglion parenchyma revealed a content (per unit area) of 40.9 +/- 8.41 NADPH-d-positive neurons and 14.42 +/- 6.7 intensely AChE-positive neurons. In the aged rats the staining intensity was unchanged, but reductions in the numbers of cells stained for NADPH-d and AChE were not significantly different. Similar counts of TH- and NPY-immunoreactive neurons revealed values of 23.2 +/- 1.77 and 19.94 +/- 4.9, respectively, suggesting frequent co-localisation. The numbers of TH- and NPY-immunoreactive neurons were found to have decreased with age by 53% and 60%, respectively. Staining of consecutive sections revealed that those neurons which stained positively for NADPH-d did not show immunoreactivity for TH or NPY, and those neurons that were immunoreactive for TH or NPY did not contain intense NADPH-d staining. Occasional NPY-1R neurons were both TH- and NADPH-d-negative. These results suggest that NADPH-d staining is predominantly confined to the parasympathetic neuron population of the hypogastric ganglion and that it is the sympathetic neuron population alone that decreases in number with age.