NADPH-diaphorase reactivity and Fos-immunoreactivity within the ventral horn of the lumbar spinal cord of cats submitted to acute muscle inflammation induced by injection of carrageenan.

The NADPH-diaphorase activity and Fos-immunoreactivity within the ventral horn of the lumbar spinal cord were studied in cats with acute unilateral myositis following injection of carrageenan into the m.m. gastrocnemius-soleus. In carrageenan-injected cats maximum in the mean number of intensely stained NADPH-diaphorase reactive (NADPH-dr) neurons was found in lamina VII (+100%) and VIII (+33%) of the contralateral ventral horn of the L6/L7 segments as compared with control animals. The maximumal level of Fos-immunoreactivity was registered in the same laminae with ipsilateral predominance (39.3±4.6 and 7.6±0.9 cells), in comparison with the contralateral side (13.6±0.8 and 5.5±0.6 cells, respectively; P<0.05). We also visualized low-intensely stained and double labelled (Fos immunoreactive+low-intensely stained NADPH-dr) multipolar and fusiform Renshaw-like cells (RLCs) within the ventral horn on both sides of the L6/L7 segments in carrageenan-injected cats. We visualized the double labelled (Fos-ir+NADPH-dr) multipolar and fusiform Renshaw-like cells (RLCs) within the ventral horn on both sides of the L6/L7 segments in carrageenan-injected cats. A significant difference in the mean number of RLCs was recorded between the ipsi- and contralateral sides in the lamina VII (13.6±2.5 vs. 4.9±0.7 cells, respectively). We suppose that activation of inhibitory RLCs in ipsilateral lamina VII could be directed on attenuation of activation of motoneurons during muscle pain development. Our study showed that a significant contralateral increase in the number of NADPH-dr cells is accompanied by an ipsilateral increase in c-Fos expression in lamina VII. These data may suggest that NADPH-dr neurons of the contralateral ventral horn through commissural connections also involved in the maintenance of the neuronal activity associated with acute muscle inflammation. It is also hypothesized, that during acute myositis, plastic changes in the ventral horn activate the processes of disinhibition due to an increase in the number of NADPH-d-reactive neurons in the spinal gray matter.

7-Nitroindazole enhances c-Fos expression in spinal neurons in rats realizing operant movements.

The expression of c-Fos and NADPH-diaphorase reactivity (NADPH-dr) in the cervical spinal cord was studied in adult male Wistar rats that realized operant reflexes after inhibition of neuronal nitric oxide synthase. Fos-immunoreactive neurons were visualized immunohistochemically in the C6/C7 spinal segments in the control, realized operant movements animals, and/or 7-nitroindazole (7-NI) injected rats. The mean numbers of immunoreactive interneurons and motoneurons (per section) were significantly greater in the Nucleus proprius (+240%) and motor nuclei (+600%) in rats of the 7-NI-pretreated and operant reflex realized group than in the isolated operant reflex realized group. Our study showed intensive staining of NADPH-dr axon terminals on the somata and initial parts of dendrites of motoneurons in experimental rats when the disodium salt of malic acid was added to the staining solution. Suppression of NO release is associated with potentiation of neuronal activation induced by descending supraspinal and proprioceptive signaling within the spinal cord.

7-Nitroindazole potentiates c-fos expression induced by muscle tendon vibration in the spinal cord.

INTRODUCTION: Expression of c-fos initiated by muscle proprioceptive signaling was studied in rats after inhibition of neuronal nitric oxide synthase (nNOS) with administration of 7-nitroindazole (7-NI). METHODS: Fos-immunoreactive (Fos-ir) neurons were visualized immunohistochemically in the lumbar cord after vibration of the Achilles tendon and/or 7-NI systemic injections. RESULTS: The total number of Fos-ir interneurons and motoneurons (per slice) was significantly greater in the 7-NI-pretreated and tendon-vibrated (7-NI + Tv) group than in the isolated tendon vibration group (Tv group). The greatest increases in the number of Fos-ir neurons were found in the L4 (+100%) and L5 (+105%) segments (P < 0.05). CONCLUSIONS: Suppression of NO release after introduction of 7-NI was associated with potentiation of Fos immunoreactivity induced by muscle proprioceptive signaling within distinctive regions of the spinal cord.

Coupling of c-fos expression in the spinal cord and amygdala induced by dorsal neck muscles fatigue.

c-fos gene expression in the cervical spinal cord and amygdala was examined in anaesthetized rats following muscle fatigue caused by intermittent high-rate (100 s(-1)) electrical stimulation of the dorsal neck muscles (m. trapezius and m. splenius). Fatigue-related increases in c-fos expression were observed on the stimulated muscle side in the cervical C2-C4 (layers 1, 3-5, 7 and 10) spinal segments, bilaterally in the lumbar L4-L6 (layer 1) segments and in contralateral central (Ce), medial (Me), and basomedial (BM) amygdaloid nuclei. A scarce number of staining cells were found within lateral and basolateral nuclei. The rostro-caudal extent of c-fos expression in the spinal cord supports functional coupling of the cervical and lumbar regions during the neck muscle fatigue development. The distinct c-fos expression in the Ce and Me amygdaloid nuclei suggests that they may contribute to mediating the neck muscle fatigue-related nociception, autonomic and behavioural responses.

Acute muscle inflammation enhances the monosynaptic reflexes and c-fos expression in the feline spinal cord.

The aim of this research was to study the changes of the motor reflex activity (monosynaptic reflex (MSR) of the flexor and extensor muscles) and Fos immunoreactivity in lumbo-sacral spinal cord after acute induced myositis of m. gastrocnemius-soleus (GS). The experiments were carried out on ischaemic decerebrated, spinalized in C1 cats. After infiltration of the GS muscle with carrageenan (2%) MSRs of flexors and extensors showed a significant increase in amplitude +127+/-24.5% and +155+/-28.5%, respectively, p<0.05. The exposed effect was initiated within 30 min and achieved a maximum 2.8h after the intramuscular injections of carrageenan. After analysis of dynamics of the MSRs, animals were perfused and c-fos expression in the spinal segments L6-S1 was evaluated. In comparison to sham-operated animals, the number of Fos-immunoreactive (Fos-ir) cells was noticeably increased in the lumbar cord of cats with carrageenan-induced myositis. The labeled cells were concentrated in the ipsilateral laminae I/II, neck of the dorsal horn (V/VI) and intermediate zone (VII), however, clear predominance of their concentration was found in the deep laminae. The effect of muscle inflammation was also expressed as a significant decline in the number of NADPH-d-reactive cells (p<0.05) in ipsilateral laminae I/II of L6/L7. The results show that the input from acutely inflamed muscles may induce an increase of the reflex responsiveness of flexors and extensors which is not mediated via the gamma-spindle-loop and which coincides with a significant increase in c-fos expression in the deep laminae of the lumbar spinal cord.

Capsaicin-induced effects on c-fos expression and NADPH-diaphorase activity in the feline spinal cord.

The distribution of c-fos expression and NADPH-diaphorase reactivity in the cervical and lumbar segments after stimulation of the vanilloid receptors in the dorsal neck muscles with capsaicin was studied in cats anaesthetized with alpha-chloralose. After the unilateral intramuscular injection of capsaicin, the mean number of Fos-immunoreactive neurons detected with an avidin-biotin-peroxidase technique was significantly increased in the superficial laminae (I), neck of the dorsal horn (V), and area around the central canal (VII) within both the cervical and lumbar spinal cord. Most Fos-immunoreactive neurons in the cervical spinal cord were giant and small cells. The widespread distribution of Fos-immunoreactive cells throughout the cervical cord within the intermediate zone (VII) coincided with the sites of localization of last-order premotor interneurons and cells of origin of inter-segmental crossed and uncrossed descending propriospinal pathways to the lumbar spinal cord. Fos-immunoreactive neurons were co-distributed with nitric oxide-generating cells at both levels of the spinal cord, although the double-labeled cells were not observed. In conclusion, the analysis of c-fos expression and NADPH-diaphorase reactivity shows that stimulation of vanilloid receptors in the neck muscles can initiate distinctive neuronal plasticity in the cervical (C1-C8) and lumbar (L1-L7) segments, and confirms the anatomical and functional coupling of both regions during processing of nociceptive signals from the dorsal neck muscles.

NO-generating neurons in the medullary cardiovascular centers of rodents and carnivores.

The aim of the study was to characterize the species-related differences in the distribution of nitric oxide synthase (NOS)-containing neurons in rodents and carnivores medullary cardiovascular centers that take part in regulation of the sympathetic or parasympathetic drives. The order of the mean number of NOS-containing neurons in the dorsomedial and ventrolateral medulla (per section) in different animals was as follows: dog>cat>rat. Although the density of the positive cells in the both regions was changed in the following sequence: rat=cat>dog. Within the dorsal motor nucleus of vagus significant exceeding of the mean number and density of positive cells (preganglionic vagal neurons) in dog were found. Differences in the distribution of NO-generating neurons in the medullary cardiovascular centers and the heterogeneity in the basal level of NO release may contribute to the distinctive alterations of the hemodynamic reactions in the studied species after administration of NOS inhibitors.

NADPH-diaphorase activity and c-fos expression in medullary neurons after fatiguing stimulation of hindlimb muscles in the rat.

In anaesthetised rats, Fos-immunoreactive and NADPH-diaphorase-positive neurons in the medulla and, for comparison, in the spinal cord were studied after fatiguing stimulation of the hindlimb muscles. Following both direct muscle stimulation and L5 ventral root stimulation, fatigue-related c-fos gene expression was most prominent in the dorsal horn of the ipsilateral L2-L5 segments and within the ipsilateral nucleus tractus solitarius, the caudoventrolateral and rostroventrolateral reticular nuclei, and the intermediate reticular nucleus at levels of -14.3 and -13.8 mm, and contralaterally at -13.2 mm caudal to the bregma. The order of intensity of c-fos expression was as follows: nucleus tractus solitarius>caudoventrolateral and rostroventrolateral reticular nuclei>intermediate reticular nucleus>lateral paragigantocellular nucleus. NADPH-diaphorase reactivity was changed in the following sequence: NTS>intermediate reticular nucleus lateral paragigantocellular nucleus>rostroventrolateral reticular nucleus. Fos-immunoreactive neurons were codistributed with NADPH-diaphorase-reactive cells within the dorsomedial and ventrolateral medulla, and double-staining neurons were found in the nucleus tractus solitarius, intermediate reticular nucleus and lateral paragigantocellular nucleus. The patterns of distribution of c-fos expression and NADPH-diaphorase reactivity show that afferent signals arising from fatiguing muscles may activate spinal and medullary neurons which are involved in nociceptive and cardiovascular reflex pathways. The functional role of nitric oxide (NO) in the generation of cardiovascular and somatosensory responses in the medulla during fatigue of skeletal muscles is discussed.