Histochemical changes of substance P, FRAP, serotonin and succinic dehydrogenase in the spinal cord of rats with adjuvant arthritis

Various histochemical changes were found in spinal segments L4-L5 of rats with adjuvant arthritis, predominantly 30 days after inoculation. A slight to marked increase of substance P immunoreactivity occurred in laminae I, II and X. FRAP activity was enhanced in lamina II. Serotonin immunoreactivity was heavier in laminae I, VIII and IX in a few animals. The intensity of the histoenzymological reaction for succinic dehydrogenase increased in certain laminae VIII and X neurons. At day 15 of the disease the increase of substance P and FRAP activities was chiefly restricted to the medial portion of the superficial dorsal horn. There was a significant positive correlation between the scratching behaviour of arthritic rats and the substance P immunoreactivity in laminae X and I. If one accepts that scratching is pain-related, the data are consistent with a possible role of substance P in the chronic pain associated with adjuvant arthritis. They leave undetermined the significance of the other histochemical changes.     

Effect of intrathecal injection of pertussis toxin on substance P, norepinephrine and serotonin contents in various neural structures of arthritic rats.

The possible influence of spinal receptors coupled to Gi/Go regulatory proteins on chronic pain adaptive processes of neural tissues was investigated in normal and arthritic rats. Pain-suffering animals showed an enhanced immunoreactivity to substance P (ir-SP) in the lumbar spinal cord, pons-medulla oblongata region and thalamus. Norepinephrine (NE) levels were increased in the spinal cord, while serotonin (5-HT) was elevated in both spinal cord and midbrain. The intrathecal injection of 1 micrograms pertussis toxin 6 days before sacrifice of rats produced in these arthritic animals a pronounced reduction of ir-SP in the pons-medulla, midbrain and thalamus, but not in the spinal cord. The level of 5-HT was diminished in dorsal spinal cord and midbrain, whereas NE appeared unchanged. In contrast, the toxin only reduced ir-SP of normal rats in the midbrain, without altering the levels of NE or 5-HT, in all the areas analysed. These results suggest the involvement of certain spinal receptors coupled to Gi/Go transducer proteins in processes leading to the elevation of ir-SP and 5-HT in various neural structures of arthritic rats.     

Intra-articular injections of high-molecular-weight hyaluronic acid have biphasic effects on joint inflammation and destruction in rat antigen-induced arthritis

To assess the potential use of hyaluronic acid (HA) as adjuvant therapy in rheumatoid arthritis, the anti-inflammatory and chondroprotective effects of HA were analysed in experimental rat antigen-induced arthritis (AIA). Lewis rats with AIA were subjected to short-term (days 1 and 8, n = 10) or long-term (days 1, 8, 15 and 22, n = 10) intra-articular treatment with microbially manufactured, high-molecular-weight HA (molecular weight, 1.7 x 10(6) Da; 0.5 mg/dose). In both tests, 10 buffer-treated AIA rats served as arthritic controls and six healthy animals served as normal controls. Arthritis was monitored by weekly assessment of joint swelling and histological evaluation in the short-term test (day 8) and in the long-term test (day 29). Safranin O staining was employed to detect proteoglycan loss from the epiphyseal growth plate and the articular cartilage of the arthritic knee joint. Serum levels of IL-6, tumour necrosis factor alpha and glycosaminoglycans were measured by ELISA/kit systems (days 8 and 29). HA treatment did not significantly influence AIA in the short-term test (days 1 and 8) but did suppress early chronic AIA (day 15, P < 0.05); however, HA treatment tended to aggravate chronic AIA in the long-term test (day 29). HA completely prevented proteoglycan loss from the epiphyseal growth plate and articular cartilage on day 8, but induced proteoglycan loss from the epiphyseal growth plate on day 29. Similarly, HA inhibited the histological signs of acute inflammation and cartilage damage in the short-term test, but augmented acute and chronic inflammation as well as cartilage damage in the long-term test. Serum levels of IL-6, tumour necrosis factor alpha, and glycosaminoglycans were not influenced by HA. Local therapeutic effects of HA in AIA are clearly biphasic, with inhibition of inflammation and cartilage damage in the early chronic phase but with promotion of joint swelling, inflammation and cartilage damage in the late chronic phase.     

Further evidence validating adjuvant arthritis as an experimental model of chronic pain in the rat

The experiments described here were aimed at further validating adjuvant arthritis as an animal model of chronic pain. It was found that the relative oral intake of a 0.008 mg/ml solution of fentanyl was higher in arthritic than in normal control rats; this difference was predicted by the notion that the analgesic effect of a substance may reinforce its intake in animals exposed to pain, more so than in normal pain-free animals. It was also found that body weight decreases and that vocalizations of aggregated rats increase as a result of the challenge; these effects suggest that the vegetative signs and the behavioral irritability which are characteristic of chronic pain in humans, also occur in arthritic animals. The pain which thus seems to be associated with adjuvant arthritis was estimated to have its onset on days 10–11, to peak on days 18–21, and to terminate on days 35–40 after inoculation with $\text{Mycobacterium butyricum}$.     

Quantitative analysis of the ultrasonic vocalization responses elicited in adjuvant-induced arthritic rats for screening analgesic drugs

Adjuvant-induced arthritic (AIA) rats have been developed as a chronic pain model to evaluate the effects of analgesic drugs. The purpose of the present study was to examine whether there is dose-dependent inhibition of the emission of ultrasonic vocalization (USV) responses by analgesic drugs in AIA rats. It was demonstrated that morphine (1.25-5.0 mg/kg, s.c.) and ketoprofen (2.5-10.0 mg/kg, s.c.) dose-dependently inhibit USV responses. These results suggest that the USV responses elicited in AIA rats are useful for the quantitative evaluation of analgesic drugs.     

Substance P-like immunoreactivity in the nervous system of hydra

Summary Using immunocytochemistry we find substance P-like material in nerve cells of hydra. These nerve cells are situated in the ectoderm of the basal disk and tentacles. Radioimmunoassay of hydra extracts gives dilution curves parallel to that of synthetic substance P, from which it can be calculated that one animal contains at least 0.6 fmol substance P-like immunoreactivity. After chromatography on Biogel P-100, the substance P-like immunoreactivity elutes as a peak in the void volume and a peak at the position of synthetic substance P.     

反复电针对慢性痛的累加治疗作用及其机制研究

本研究从基础和临床两方面观察了反复电针对慢性痛的累加治疗作用,并结合疼痛患者及慢性痛动物模型中几种神经肽的放射免疫测定及相应受体拮抗剂的药理学研究结果,探讨了产生累加效应的可能机制。结果表明,在临床脊髓损伤性痉挛患者,１００Ｈｚ穴位体表电刺激有效地缓解痉挛并有累加效应;在临床慢性痛患者,２／１５Ｈｚ变频ＴＥＮＳ刺激有效地治疗疼痛并具有累加效应。在关节炎模型大鼠,电针刺激能产生明显的镇痛并具有累加效     

Substance P-like immunoreactivity in the nervous system of hydra

Using immunocytochemistry we find substance P-like material in nerve cells of hydra. These nerve cells are situated in the ectoderm of the basal disk and tentacles. Radioimmunoassay of hydra extracts gives dilution curves parallel to that of synthetic substance P, from which it can be calculated that one animal contains at least 0.6 fmol substance P-like immunoreactivity. After chromatography on Biogel P-100, the substance P-like immunoreactivity elutes as a peak in the void volume and a peak at the position of synthetic substance P.     

Ultrasonic vocalization response elicited in adjuvant-induced arthritic rats as a useful method for evaluating analgesic drugs.

Adjuvant-induced arthritic (AIA) rats develop a severe chronic polyarthritis which shares some features in common with human rheumatoid arthritis. The purpose of the present study was to examine whether AIA rats emit ultrasonic vocalizations (USVs) when they are confronted with a healthy 'stimulus rat' in social interactions. We also examined the effects of three analgesic drugs (piroxicam, rofecoxib and ketoprofen) on USV responses using the same paradigm. In social interactions, AIA rats and intact controls emitted USVs in the 22-28 kHz range. Vocalization activities were significantly higher in AIA rats than those in intact controls. Moreover, the USVs of AIA rats were significantly inhibited by the three analgesic drugs. These results suggest that the USV responses elicited in AIA rats are useful for the evaluation of analgesic drugs.     

Localisation of substance P-like immunoreactivity in the ciliary ganglia of monkey (Macaca fascicularis) and cat: a light- and electron-microscopic study

The present study describes substance P-like immunoreactivity in the ciliary ganglia of monkey (Macaca fascicularis) and cat. About 60% of neurons in the monkey ciliary ganglion and 40% in the cat ciliary ganglion were substance P-like immunoreactive, ranging from faint to moderate staining. Substance P-like immunoreactivity was located in cell bodies, dendritic profiles and axons. In the monkey, substance P-like immunoreactive pericellular arborisations were associated with about 0.5%–3% of the ganglion cells, which were either negatively, faintly or moderately stained. An electron-microscopic study demonstrated the presence of either substance P-like immunoreactive positive or negative axon terminals synapsing or closely associated with positive dendritic profiles in both the monkey and cat ciliary ganglia. The results suggest that substance P plays an important role in the ciliary ganglion, perhaps as a modulator or transmitter.     

Neurokinin A-like immunoreactivity in rat primary sensory neurons; coexistence with substance P

Summary Rat spinal cord, dorsal root ganglia and skin were investigated employing immunohistochemical technique with specific antisera to neurokinin A and substance P. Neurokinin A-like immunoreactivity was detected in the spinal dorsal horn and skin with a similar distribution pattern as that of substance P-like immunoreactivity. After dorsal root transection a parallell decrease of neurokinin A and substance P-like immunoreactivity was observed in the dorsal horn. Using colchicine pretreatment a population of neurokinin A positive cell bodies was seen in the dorsal root ganglia, and by comparison of consecutive sections of the same cells stained for substance P it was revealed that these neurons also display substance P-like immunoreactivity. However, substance P-, but not neurokinin A-, immunoreactive cells were also observed. It is concluded that neurokinin A- and substance P-like immunoreactivity coexist in a population of rat primary sensory neurons.     

Neurokinin A-like immunoreactivity in rat primary sensory neurons; coexistence with substance P

Rat spinal cord, dorsal root ganglia and skin were investigated employing immunohistochemical technique with specific antisera to neurokinin A and substance P. Neurokinin A-like immunoreactivity was detected in the spinal dorsal horn and skin with a similar distribution pattern as that of substance P-like immunoreactivity. After dorsal root transection a parallel decrease of neurokinin A and substance P-like immunoreactivity was observed in the dorsal horn. Using colchicine pretreatment a population of neurokinin A positive cell bodies was seen in the dorsal root ganglia, and by comparison of consecutive sections of the same cells stained for substance P it was revealed that these neurons also display substance P-like immunoreactivity. However, substance P-, but not neurokinin A-, immunoreactive cells were also observed. It is concluded that neurokinin A- and substance P-like immunoreactivity coexist in a population of rat primary sensory neurons.     

Combined retrograde tracing and immunohistochemistry of trigeminal ganglion neurons projecting to gingiva or tooth pulps in the lower jaw of the cichlid Tilapia mariae

Rat trigeminal ganglion neurons projecting to the oral mucosa or to tooth pulps have different cell diameters and contain different chemical markers. In the present paper we examine whether trigeminal ganglion neurons sending axons to gingiva or tooth pulps in the lower jaw of the cichlid Tilapia mariae differ in a similar way. Retrograde tracing with fluorescent latex microspheres revealed labelled gingival and pulpal neurons in the caudal part of the trigeminal ganglion. The gingival neurons had a unimodal size distribution (peak 11 μm; range 8–14 μm) and the pulpal neurons exhibited a bimodal size distribution (peaks 12 and 25 μm; range 10–40 μm). Immunohistochemistry revealed a calcitonin gene-related peptide-like immunoreactivity in some 40% of the gingival neurons and a substance P-like immunoreactivity in 30%. Of the small pulpal neurons about 60% exhibited a calcitonin gene-related peptide-like immunoreactivity and 15% showed a substance P-like immunoreactivity. Of the large pulpal neurons some 70% exhibited a calcitonin gene-related peptide-like immunoreactivity. These neurons did not show a substance P-like immunoreactivity. In some animals a few trigeminal ganglion neurons showed a neuropeptide Y- or a vasoactive intestinal polypeptide-like immunoreactivity. Perikarya with a tyrosine hydroxylase- or a choline acetyl transferase-like immunoreactivity were not observed. We conclude that gingiva and tooth pulps in the lower jaw of T. mariae are innervated by trigeminal ganglion neurons, the cell diameters and neuropeptide contents of which differ in a pattern similar to that in the rat. Hence, this seems to represent a conserved evolutionary pattern.     

The colocalization of substance P- and somatostatin-like peptides in neurons of the entopeduncular nucleus of rats

The colocalization of substance P-like and somatostatin-like immunoreactivity in cell bodies was investigated in the entopeduncular nucleus of rats by a double immunofluorescence method using species specific antibodies. Most of the substance P-like immunoreactive cells were also positive to somatostatin and mainly seen in the rostral to middle region of the entopeduncular nucleus. Therefore it is suggested that double-labeled neurons in the entopeduncular nucleus project to the lateral habenular nucleus which is involved in the limbic system, since the rostral portion of the entopeduncular nucleus has been shown to project to the lateral habenular nucleus.     

Immunochemical evidence for substance P-like peptide in tissues of the earthworm Lumbricus terrestris: action on intestinal contraction

The presence of a substance P-like peptide in intestinal and body wall tissues, ventral nerve fiber and seminal vesicles of the earthworm Lumbricus terrestris has been demonstrated by means of a radioimmunoassay technique. The greatest substance P-like immunoreactivity was measured in intestinal tissues where it stimulates the rate of spontaneous contraction. This effect is inhibited by the substance P antagonist (D-pro2, D-trp7,9)-SP suggesting a possible involvement of receptor mechanisms. Dual localization of substance P-like immunoreactivity in earthworm intestinal and nerve tissues follows the pattern observed of peptidal hormones in vertebrates which are common to both endocrine and non-endocrine tissues.     

Intimate relationship between interstitial cells of Cajal and enteric nerves in the guinea-pig small intestine

Recent studies have suggested that enteric inhibitory neurotransmission is mediated via interstitial cells of Cajal in some gastrointestinal tissues. This study describes the physical relationships between enteric neurons and interstitial cells of Cajal in the deep muscular plexus (IC-DMP) of the guinea-pig small intestine. c-Kit and vimentin were colocalized in the cell bodies and fine cellular processes of interstitial cells of the deep muscular plexus. Anti-vimentin antibodies were subsequently used to examine the relationships of interstitial cells with inhibitory motor neurons (as identified by nitric oxide synthase-like immunoreactivity) and excitatory motor neurons (using substance P-like immunoreactivity). Neurons with nitric oxide synthase- and substance P-like immunoreactivities were closely associated with the cell bodies of interstitial cells and ramified along their processes for distances greater than 300 7m. With transmission electron microscopy, we noted close relationships between interstitial cells and the nitric oxide synthase- and substance P-like immunoreactive axonal varicosities. Varicosities of nitric oxide synthase and substance P neurons were found as close as 20 and 25 nm from interstitial cells, respectively. Specialized junctions with increased electron density of pre- and postsynaptic membranes were observed at close contact points between nitric oxide synthase- and substance P-like immunoreactive neurons and interstitial cells. Close structural relationships (approximately 25 nm) were also occasionally observed between either nitric oxide synthase- and substance P-like immunoreactive varicosities and smooth muscle cells of the outer circular muscle layer. The data suggest that interstitial cells in the deep muscle plexus are heavily innervated by excitatory and inhibitory enteric motor neurons. Thus, these interstitial cells may provide an important, but probably not exclusive, pathway for nerve-muscle communication in the small intestine.     

Elevation of Substance P-like Immunoreactivity in Rat Central Nervous System by Protease Inhibitors

Abstract: Several substance P-rich areas in rat CNS had increased levels of substance P-like immunoreactivity following the intraventricular injection of the protease inhibitors SQ 20881, SQ 14225, and leupeptin. There were significant differences in response patterns from region to region, possibly on account of an interaction of anatomical, biochemical, or physiological variables. Although the compound SQ 14225 appeared to be the most potent of the inhibitors examined, it had no apparent effect on CNS substance P-like immunoreactivity when administered peripherally.     

Differential effects of treadmill exercise in early and chronic diabetic stages on parvalbumin immunoreactivity in the hippocampus of a rat model of type 2 diabetes

In the present study, we investigated the effects of treadmill exercise in early and chronic diabetic stages on parvalbumin (PV) immunoreactivity in the subgranular zone of the dentate gyrus of Zucker diabetic fatty (ZDF) and its lean control rats (ZLC). To investigate the effects, ZLC and ZDF rats at 6 or 23 weeks of age were put on a treadmill with or without running for 1 h/day/5 consecutive days at 16–22 m/min for 5 weeks or 12–16 m/min for 7 weeks, respectively. Physical exercise in pre-diabetic rats prevented onset of diabetes, while exercise in rats at chronic diabetic stage significantly reduced blood glucose levels. In addition, physical exercise in the pre-diabetic rats significantly increased PV immunoreactive fibers in the strata oriens and radiatum of the CA1-3 region and in the polymorphic and molecular layers of the dentate gyrus compared to that in sedentary controls. However, in rats at chronic stages, PV immunoreactivity was slightly increased in the CA1-3 region as well as in the dentate gyrus compared to that in the sedentary controls. These results suggest that physical exercise has differential effects on blood glucose levels and PV immunoreactivity according to diabetic stages. Early exercise improves diabetic phenotype and PV immunoreactive fibers in the rat hippocampus.     

Intrathecal pertussis toxin attenuates the morphine withdrawal syndrome in normal but not in arthritic rats

The effect of intrathecal pertussis toxin on morphine dependence was studied in rats suffering from chronic pain (Freund's adjuvant-induced arthritis). Animals were rendered tolerant-dependent by subcutaneous implantation of 3 pellets of 75 mg morphine base each. In both, normal and arthritic animals, 1 microgram pertussis toxin reduced the analgesia induced by morphine in the tail-flick test. Naloxone (1 mg/kg, s.c.) precipitated a withdrawal syndrome in arthritic animals that was milder in respect to the one produced in normal rats. Pretreatment with pertussis toxin significantly diminished the incidence of withdrawal signs such as jumps, squeak on touch, chattering, ptosis, body shakes and diarrhoea in tolerant-dependent normal rats, while this effect could not be observed in animals suffering from chronic pain. This differential activity of the toxin could be due to the altered tonus of certain neurotransmitter systems that accompanies the chronic situation of pain.