An Immunohistochemical Study on the Presence of Nitric Oxide Synthase Isoforms (nNOS, iNOS, eNOS) in the Spinal Cord and Nodose Ganglion of Rats Receiving Ionising Gamma Radiation to their Liver.

INTRODUCTION: This study determined the presence of nitric oxide synthesis isoforms (nNOS, iNOS, and eNOS) in thoracic spinal cord segments and nodose ganglia of rats with gamma-irradiated livers. MATERIAL AND METHODS: Male rats (n = 32) were divided into equal groups A, B, C, and D. In group A, the controls, no radiation was applied, while groups B, C, and D received 10 Gy of ionising gamma radiation. The rats of group B were euthanized at the end of the first day (d1), those of group C on the second day (d2), and those of group D on the third day (d3). The liver, spinal cord segments, and nodose ganglion tissues were dissected and fixed, and the liver sections were examined histopathologically. The other tissues were observed through a light microscope. RESULTS: Regeneration occurred at the end of d3 in hepatocytes which were radiation-damaged at the end of d1 and d2. On d1, some nNOS-positive staining was found in the neuronal cells of laminae I-III of the spinal cord and in neurons of the nodose ganglion, and on d3, some staining was observed in lamina X of the spinal cord, while none of note was in the nodose ganglion. Dense iNOS-positive staining was seen on d1 in the ependymal cells of the spinal cord and in the glial cells of the nodose ganglion, and on d3, there was still considerable iNOS staining in both tissues. There was clear eNOS-positive staining in the capillary endothelial cells of the spinal cord and light diffuse cytoplasmic staining in the neurons of the nodose ganglion on d1, and on d3, intense eNOS-positive staining was visible in several endothelial cells of the spinal cord, while light nuclear staining was recognised in the neurons of the nodose ganglion. CONCLUSION: The nNOS, iNOS, and eNOS isoforms are activated in the spinal cord and nodose ganglion of rats after ionising radiation insult to the liver.

Three-dimensional computed tomographic examination of pelvic cavity in Van Cats and its morphometric investigation.

This study was carried out to determine the morphometric values of the pelvic cavity, the cranial and caudal pelvic aperture by three-dimensional (3D) reconstruction of the images obtained by computed tomography in adult Van Cats and to show the differences between these indicators in both genders. A total of 16 adult Van Cats, eight male and eight female, were used in the study. Animals were anesthetized with ketamine-xylazine combination and were scanned by computerized tomography. Images of the pelvic cavity were obtained. Three-dimensional reconstruction of the pelvis was performed with three-dimensional modelling program. Measurements of the diameters of the pelvic cavity, the cranial and caudal pelvic aperture were calculated from these images, and statistical analyzes were performed. Three-dimensional modelling of the pelvic cavity of Van Cats were examined between males and females. The results showed that there were significant (p < .05) differences between measurement of the pelvic inclination, the medial transverse diameter, the caudal transverse diameter and the angle between ischiatic arch. In addition, the correlations between age and weight were found statistically significant (p < .05) with the dorsal transverse diameter, the intermediate transverse diameter and the caudal transverse diameter in males. On the other hand, the correlations of age were statistically significant (p < .01) with the cranial pelvic aperture and the caudal pelvic aperture in females. In conclusion, some biometric values of pelvic cavity were observed to be statistically different in Van Cats.

A semi-quantitative analysis of Fos expression by mustard oil.

In the present study, a semi-quantitative analysis of Fos expression by mustard oil was performed. For this purpose, mustard oil was applied to the skin of the right hind foot of Wistar rats at various concentrations: 5%, 10%, 30%, 50%, 80% and 100% in liquid paraffin. The distribution and number of activated Fos-positive cells in the stimulated side (ipsilateral) and contralateral side of the spinal cord were investigated following the application. The ED50 of the response was also determined. The number of Fos-labelled cells gradually increased in a dose dependent manner in both sides of superficial layers (laminae I-II) of the spinal cord with increasing concentration of mustard oil. The increase between the doses was found significant in the ipsilateral superficial layers. The increase was significant in the contralateral superficial layers at concentrations above 50%. Very few Fos-labelled cells were observed around the central canal region in all concentrations. Higher doses of the mustard oil did not increase the number of activated cells in the deeper layers. However, the expression in the deeper layers (laminae III-X) does not show a consistent trend. Also, none of the concentrations used produced labelling in neurons of the deep ventral horn neurons or in motor neurons. Forty percent (40%) of mustard oil gave an approximately 1/2 maximum response i.e. an approximate ED 50. This may be important for studies using intrathecal application of antagonist following the mustard oil activation of skin nerve fibres.

Multiple neurotransmitter receptors contribute to the spinal Fos expression.

The aim of this study is to identify the receptors which could potentially mediate the activation of c-Fos. Therefore, the effects of neurotransmitter receptor agonists in the activation of c-Fos in spinal neurons were studied by intrathecal injection of excitatory amino acid (EAA) receptor agonists: N-Methyl-D-Aspartate (NMDA), (S)-alpha-Amino-3-Hydroxy-5-Methyl-4-Isoxazolepropionic acid (AMPA), 2-Carboxyl-3-carboxymethyl-4-isopropenylpyMidine (Kainic acid, KA), (1S-3R)-1-Aminocyclopentane-1, 3-dicarboxylacid (ACPD), and substance-P receptor (neurokinin-1) agonist, [Sar9, Met (O2)11] SP (SarMet-SP). All drugs tested activated the production of c-Fos in spinal dorsal horn neurons. AMPA was found as the most potent agonist tested producing market production of c-Fos particularly in neurons of lamina II at doses of 10 pM per 10-microl injection. At this dose, other agonists were relatively ineffective. At higher doses, AMPA significantly increased the activated cells. NMDA significantly increased c-Fos production to a marked extent only at doses above 10 nM per 10-microl injection. KA and ACPD were least potent of the excitatory amino acid agonists. Injection of SarMet-SP at doses of 1 nM activated Fos selectively in neurons of lamina I. A dose-dependent increase in number of c-Fos-positive cells was observed for AMPA, KA, ACPD, and SarMet-SP, whereas NMDA gave a very strong expression after a high dose with no dose dependency. These finding suggest that multiple neurotransmitter receptors lead to c-Fos production in spinal neurons.

NMDA and AMPA/KA receptors are involved in the c-Fos expression following mustard oil activation of C-fibres.

To examine whether mustard oil application to the skin activated c-Fos via glutamate receptors, a competitive NMDA receptor antagonist, 3-(2-carboxpiperazin-4-yl)propyl-1-phospionic acid (CPP), a selective AMPA/KA receptor antagonist, 6-cyano-2,3-dihydroxy-7-nitro-quinoxaline (CNQX), or both, were used intrathecally 10 min prior to noxious stimulation. Application of mustard oil to left hind foot of the vehicle-injected animals produced c-Fos expression mainly in superficial laminae (laminae I-II) of the dorsal horn on the side ipsilateral to the stimulation. CPP significantly reduced the number of c-Fos-positive nuclei in superficial laminae. But significant reduction of c-Fos expression by CNQX was seen in deeper laminae (laminae III-X). Administration of both CPP and CNQX extensively reduced the number of c-Fos-positive cells in both superficial and deeper laminae. However, they did not greatly change the number of c-Fos-positive cells in lamina I. This experiment revealed that NMDA and AMPA/KA receptors contribute to the mustard oil-induced c-Fos expression in the spinal cord. These data also suggest that other neurotransmitter receptors might be involved in the activation produced by algesic chemical activation of C-fibre primary afferents.

Neuronal nitric oxide synthase immunoreactivity in ependymal cells during early postnatal development.

Neuronal nitric oxide synthase (nNOS) immunoreactivity was observed in ependymal cell layer of the central canal of spinal cord of neonatal rats (2-20 days old). Neuronal nitric oxide synthase immunoreactivity was present in postnatal day 2 and this immunoreactivity gradually disappeared by postnatal day 16. The progressive decrease in nNOS staining with the increasing postnatal age may suggest that nNOS staining paralleled the maturation of the central canal and may also suggest that nNOS activity plays a role in the development of the ependymal cells.