Although guanethidine (GUA) was used in the past as a drug to suppress hyperactivity of the sympathetic nerve fibers, there are no available data concerning the possible action of this substance on the sensory component of the peripheral nervous system supplying the urinary bladder. Thus, the present study was aimed at disclosing the influence of intravesically instilled GUA on the distribution, relative frequency, and chemical coding of dorsal root ganglion neurons associated with the porcine urinary bladder. The investigated sensory neurons were visualized with a retrograde tracing method using Fast Blue (FB), while their chemical profile was disclosed with single-labeling immunohistochemistry using antibodies against substance P (SP), calcitonin gene-related peptide (CGRP), pituitary adenylate cyclase activating polypeptide (PACAP), galanin (GAL), neuronal nitric oxide synthase (nNOS), somatostatin (SOM), and calbindin (CB). After GUA treatment, a slight decrease in the number of FB⁺ neurons containing SP was observed when compared with untreated animals (34.6 ± 6.5% vs. 45.6 ± 1.3%), while the number of retrogradely traced cells immunolabeled for GAL, nNOS, and CB distinctly increased (12.3 ± 1.0% vs. 7.4 ± 0.6%, 11.9 ± 0.6% vs. 5.4 ± 0.5% and 8.6 ± 0.5% vs. 2.7 ± 0.4%, respectively). However, administration of GUA did not change the number of FB⁺ neurons containing CGRP, PACAP, or SOM. The present study provides evidence that GUA significantly modifies the sensory innervation of the porcine urinary bladder wall and thus may be considered a potential tool for studying the plasticity of this subdivision of the bladder innervation.

Phoenixin-14 (PNX), initially discovered in the rat hypothalamus, was also detected in dorsal root ganglion (DRG) cells, where its involvement in the regulation of pain and/or itch sensation was suggested. However, there is a lack of data not only on its distribution in DRGs along individual segments of the spinal cord, but also on the pattern(s) of its co-occurrence with other sensory neurotransmitters. To fill the above-mentioned gap and expand our knowledge about the occurrence of PNX in mammalian species other than rodents, this study examined (i) the pattern(s) of PNX occurrence in DRG neurons of subsequent neuromeres along the porcine spinal cord, (ii) their intraganglionic distribution and (iii) the pattern(s) of PNX co-occurrence with other biologically active agents. PNX was found in approximately 20% of all nerve cells of each DRG examined; the largest subpopulation of PNX-positive (PNX⁺) cells were small-diameter neurons, accounting for 74% of all PNX-positive neurons found. PNX⁺ neurons also co-contained calcitonin gene-related peptide (CGRP; 96.1%), substance P (SP; 88.5%), nitric oxide synthase (nNOS; 52.1%), galanin (GAL; 20.7%), calretinin (CRT; 10%), pituitary adenylate cyclase-activating polypeptide (PACAP; 7.4%), cocaine and amphetamine related transcript (CART; 5.1%) or somatostatin (SOM; 4.7%). Although the exact function of PNX in DRGs is not yet known, the high degree of co-localization of this peptide with the main nociceptive transmitters SP and CGRP may suggests its function in modulation of pain transmission.

The study was designed to examine the distribution and chemical coding of somatostatin-immunoreactive (SOM-IR) nerve fibers supplying the urinary bladder wall and to establish the distribution and immunohistochemical characteristics of the subpopulation of paracervical ganglion (PCG) SOM-IR neurons projecting to this organ in female pigs. The PCG-urinary bladder projecting neurons (PCG-UBPN) were visualized with retrograde neuronal tracer Fast Blue (FB). Double-labeling immunohistochemistry performed on cryostat sections from the urinary bladder wall revealed that the greatest density of SOM-IR nerve fibers was found in the muscle layer and around blood vessels, a moderate number of these nerve terminals supplied the submucosa and only single SOM-IR axons were encountered beneath the urothelium. In all the investigated sections the vast majority of SOM-IR nerve fibers were immunopositive to vesicular acetylcholine transporter (VAChT) and many SOM-IR axons contained immunoreactivity to neuropeptide Y (NPY). Approximately 65 % of FB-positive (FB+) PCG-UBPN were immunoreactive to SOM. Moreover, PCG FB+/SOM + nerve cells were simultaneously immunoreactive to choline acetyltransferase (ChAT; 64.6 ± 0.6 %), NPY (59.7 ± 1.2 %), neuronal nitric oxide synthase (nNOS; 46.1 ± 0.7 %), vasoactive intestinal polypeptide (VIP; 29.9 ± 2.2 %), Leu⁵-enkephalin (L-ENK; 19.5 ± 6.3 %), dopamine β-hydroxylase (DβH; 14.9 ± 1.9 %) or pituitary adenylate cyclase-activating polypeptide (PACAP; 14.8 ± 2.4 %). The present study reveals the extensive expression of SOM in both the nerve fibres supplying the porcine urinary bladder wall and the PCG neurons projecting to this organ, indicating an important regulatory role of SOM in the control of the urinary bladder function.

Phoenixin-14 (PNX) - a bioactive peptide recently discovered in the rat brain, is highly conserved among many animal species: rodents (rat, mouse), pig, dog as well as humans. In rodents, PNX is expressed in areas responsible for the transmission of sensory information dorsal root ganglia (DRG) neurons, where it is present at relatively high levels and may preferentially suppress visceral pain. However, so far the presence of PNX has not been investigated in DRG in pigs, a species which, due to its anatomical and histological similarity to humans, is considered a better model for biomedical studies than rodents. The present study aimed to investigate the immunoreactivity of PNX in the DRG of the domestic pig. The collected spinal ganglia from the cervical (C), thoracic (Th), lumbar (L) and sacral (S) sections were transversely divided into serial sections of 10 μm thickness. DRG sections from each level of the spinal cord were double-labeled immunohistochemically using antibodies to PNX in a mixture with antibodies to: cocaine and amphetamine related transcript (CART), calretinin (CRT), calcytonin gene-related peptide (CGRP), galanin (GAL), nitric oxide synthase (nNOS), pituitary adenylate cyclase-acrivating polypeptide (PACAP), substance P (SP) or somatostatin (SOM), respectively. Immunohistochemical studies revealed PNX immunoreactivity in approximately 20% of nerve cells in all DRG examined, highlighting mainly the presence of the peptide in cells of small diameter (approximately 74% of all PNX-positive neurons found). Double labeling of DRG sections showed that PNX-immunopositive neurons stained also for CGRP (96.1%), SP (88.5%), nNOS (52.1%), GAL (20.7%), CRT (10.05%), PACAP (7.4%), CART (5.1%), or SOM (4.7%). Our research revealed for the first time the presence of the new peptide PNX in the sensory ganglia of the domestic pig, its co-localization with other important neurotransmitters involved in sensory transmission and its percentage distribution in ganglion domains. The exact function of PNX in DRG is not yet known, however, the high degree of co-localization of this peptide with the main nociceptive transmitters SP and CGRP may indicate its function in modulation of pain transmission.

One of most important parts of the female genital tract is the uterine cervix, both from the anatomical as well as physiological points of view. As there is currently a lack of detailed information on the presence, distribution pattern(s), and the chemical coding of phoenixin (PNX)-containing dorsal root ganglia (DRG) neurons supplying the porcine uterine cervix, this study, using combined retrograde tracing and double-immunofluorescence techniques, was aimed at analyzing (i) the distribution pattern of uterine cervix-supplying sensory neurons (UC-SNs) at the particular spinal cord levels, (ii) their intraganglionic distribution, and (iii) the patterns of PNX co-expression with other biologically active substances. UC-SNs were identified by the presence of deposits of Fast Blue (FB), in DRG of thoracic (Th10–Th15), lumbar (L1–L5) and sacral (S2–S4) spinal cord segments. FB⁺/PNX⁺ neurons constitute approximately 33% of all UC-SNs, 73% at the L, and 27% at the S neuromeres. These neurons were mainly small sized (52%), with a slightly smaller population of medium-sized cells (40%), while large-diameter cells made up the least numerous population (8%). The vast majority of FB⁺/PNX⁺ neurons simultaneously contained calcitonin gene-related peptide (CGRP; 80.9%) or substance P (SP; 77.9%); one-third of them showed immunoreactivity toward neuronal nitric oxide synthase (nNOS; 34%), while PNX⁺ UC-SNs containing pituitary adenylate cyclase-activating polypeptide (PACAP), galanin (GAL), calretinin (CRT), or somatostatin (SOM) formed significantly smaller populations (21.4%, 7.4%, 3.1%, and 0.7%, respectively). The results of the present study demonstrate the presence of PNX in DRG UC-SNs, and its co-occurrence with numerous neurotransmitters suggesting a putative role for this neuropeptide in the transmission of various types of sensory information and possible effects on the functioning of this organ in the body.

The present study was designed to establish the distribution pattern and immunohistochemical characteristics of phoenixin-immunoreactive (PNX-IR) urinary bladder afferent neurons (UB-ANs) of dorsal root ganglia (DRG) in female pigs. The sensory neurons investigated were visualized with a retrograde tracing method using Fast Blue (FB), while their chemical profile(s) were identified using double-labelling immunohistochemistry with antibodies against PNX, calcitonin gene-related peptide (CGRP), calretinin (CRT), galanin (GAL), neuronal nitric oxide synthase (nNOS), pituitary adenylate cyclase-activating polypeptide (PACAP), somatostatin (SOM) and substance P (SP). Nearly half of UB-ANs contained PNX (45%), and the majority of such encoded sensory neurons were small in size (66%). The most numerous subpopulation of FB/PNX-positive neurons were those containing SP (71%). CGRP, GAL or PACAP were observed in a smaller number of PNX-containing UB-ANs (50%, 30% or 25%, respectively), while PNX-positive sensory neurons simultaneously immunostained with nNOS, CRT or SOM constituted a small fraction of all retrogradely-traced DRG neurons (DRGs; 15%, 6.5% or 1.6%, respectively). Furthermore, the numerical analysis of neurons expressing individual antigens, performed on 10 μm-thick consecutive sections, allows us to state that studied sensory neurons can be classified as neurons “coded” either by the simultaneous presence of SP/CGRP/PACAP/GAL, SP/CGRP/PACAP/NOS, SP/CGRP/PACAP/NOS/CRT and/or SP/CGRP/GAL/PACAP, or, as a separate population, those capable of SOM synthesis (SP/CGRP/SOM/PACAP/GAL-positive neurons). The present study reveals the extensive expression of PNX in the DRGs supplying to the urinary bladder, indicating an important regulatory role of this neuropeptide in the control of physiological function(s) of this organ.