Biomedical subjects
Publications and source records attributed to J M Besson.
Pain researchers are committed to designing new experimental models as close as possible to the physiopathological conditions encountered in humans, with due concern for ethical considerations. Some current questions related to inflammation and headache are discussed, along with the approaches used to address them. These approaches are essential for a better understanding of the physiology, pharmacology, and treatment of various pain syndromes.
We have recently developed, in the rat, a model with a limited arthritic process for chronic pain studies. Intra-articular injection (0.05 ml) of complete adjuvant containing 300 micrograms Mycobacterium butyricum in the tibio-tarsal joint produces a predictable monoarthritis stable clinically and behaviourly from weeks 2 through 6 post-injection. This model appears to be a suitable alternative for the polyarthritic rat for chronic studies based on both its ethical and scientific advantages. In the present work we report results of experiments on the effects of exercise on the pain behaviour and development of arthritis in this model. A group of rats prepared with the above protocol was submitted at 2 weeks post-inoculation to mild exercise (swimming [water 37 degrees C] three times per week) increasing from 5 to 15 min during 4 weeks. As revealed by analyses of the arthritis score, the stiffness score and the mobility score, no aggravation of arthritis occurred in these rats. However the threshold for struggle in response to paw pressure was further decreased (as compared to control arthritic rats) in these animals. These results are discussed in view of observations made in human studies.
Parallel time courses of clinical and behavioural parameters and levels of plasma substance P-like immunoreactivity (SPLI-PI) were studied in arthritic rats (adjuvant induced arthritis, AIA, a chronic pain model). Acute (14 and 21 post-inoculation days,PI) and post-acute (42 days PI) phases of the syndrome were investigated. These data were compared with those obtained in a control situation (inoculation day). In a second experimental series, levels of substance P-like immunoreactivity in cerebrospinal fluid (SPLI-CSF) were determined at the same stages of AIA. In arthritic rats SPLI-PI was strongly enhanced (X4) as early as 14 days PI and remained increased (X4) at all stages studied, whereas SPLI-LCR was significantly increased (X2) only 21 days PI and returned to control levels at 42 days PI. These data suggest that SP could be distributed in two different pools, a peripheral one of inflammatory origin, and a central one which could be more specific to the chronic pain situation.
Neurons (n = 107) were recorded, in anaesthetized rats, with extracellular micropipettes, in the nucleus centralis of the amygdala (Ce). The activity of a large majority (77%) was exclusively or preferentially affected by noxious stimuli applied in any area of the body. The rest of the neurons was not activated by any stimuli (21%) or responded only to innocuous stimuli (2%). The nociceptive neurons were classified into two groups: the neurons in the first group (34% of the whole population) were activated by mechanical and thermal noxious stimuli; the neurons in the second group (43% of the whole population) were inhibited by noxious stimuli. Neurons in two groups precisely encoded thermal stimuli in the noxious range. Transcutaneous electrical stimulation suggested that the Ce nociceptive neurons were under the influence of A delta and C fibers. These results demonstrate important nociceptive input to the Ce which according to previous studies probably relays in the pontine parabrachial nucleus. In addition, they are in good agreement with the involvement of the amygdala in emotional, autonomic and/or behavioral reactions to noxious events.
Highly selective tritiated ligands and quantitative autoradiography have been used to study mu, delta and kappa binding sites in the dorsal horn of the rat spinal cord. We have measured the proportions of the 3 main types of opioid binding sites in the superficial layers of the cervical dorsal horn (laminae I and II). The proportions of mu, delta and kappa sites were 70 +/- 4%, 23 +/- 2% and 7 +/- 1%, respectively, over the whole C4-T2 extent. Similar percentages were encountered at the level of each individual segment from C4 to T2. Eight days after a unilateral dorsal rhizotomy C4-T2, dramatic decreases were seen on the ipsilateral side to the lesion by comparison to the intact side. In the C7 segment, these decreases were 76 +/- 1%, 61 +/- 1% and 53 +/- 3% for mu, delta and kappa binding sites, respectively. The C7 segment can be considered as completely deafferented, so we attribute the residual values to postsynaptic binding whereas the decrease can be attributed to a loss of the presynaptic sites. These results are discussed with respect to the contribution of pre- and postsynaptic depressive effects of opiates on the transmission of noxious messages at the level of the dorsal horn.
The effect of administering low doses (0.5-1.5 micrograms) of the mu-opioid receptor agonist fentanyl into the right brachial plexus sheath of the rat was examined using the vocalization threshold to paw pressure test. Both forepaws were tested in each rat. Fentanyl injected into the right brachial plexus sheath at 0.5-1.5 micrograms/kg produced a localized, dose-dependent, potent and long lasting antinociceptive effect, as gauged on the right forepaw. At the lower dose used (0.5 microgram/kg of fentanyl), the antinociceptive effect was restricted to the right forepaw and lasted for more than 2 h. Increasing doses of fentanyl (1 and 1.5 micrograms/kg) induced potent effects, lasting up to 5-6 h or even longer. In complete contrast, fentanyl administered i.v. at the dose of 1 microgram/kg had a very transient effect, only lasting up to 25 min. The results of injection of low doses of the opioid antagonist naloxone when administered either i.v. or locally into the paw, on the effect of fentanyl suggest the involvement of a peripheral site of action of the opioid. The present findings suggest that, as already observed in patients in clinical situations, low doses of opiates delivered using this administration route may provide prolonged regional analgesia, with the potential of avoiding centrally mediated side effects.
Biochemical mapping of five different peptide-like materials--calcitonin gene-related peptide (CGRP), substance P (SP), Met5-enkephalin (ME), cholecystokinin (CCK), and dynorphin A (1-8) (DYN)--was conducted in the dorsal and ventral zones of the spinal cord at the cervical, thoracic, and lumbar levels in 3-month-old rats 10 days after unilateral dorsal rhizotomy at the cervical level (C4-T2) or after neonatal administration of capsaicin (50 mg/kg s.c.). In control rats, all peptide-like materials were more abundant in the dorsal than in the ventral zone all along the spinal cord. However, in both zones, absolute concentrations of CGRP, SP, ME, and CCK were significantly higher at the lumbar than at the cervical level. Rhizotomy-induced CGRP depletion (-85%) within the ipsilateral dorsal zone of the cervical cord was more pronounced than that due to neonatal capsaicin (-60%), a finding suggesting that this peptide is contained in both capsaicin-sensitive (mostly unmyelinated) and -insensitive (myelinated) primary afferent fibers. In contrast, similar depletions of SP (-50%) were observed after dorsal rhizotomy and neonatal capsaicin treatment, as expected from the presence of SP only in the capsaicin-sensitive small-diameter primary afferent fibers. Although the other three peptides remained unaffected all along the cord by either intervention, evidence for the existence of capsaicin-insensitive CCKergic primary afferent fibers could be inferred from the increased accumulation of CCK (together with SP and CGRP) in dorsal root ganglia ipsilateral to dorsal root sections.
1. Neurons were recorded in the parabrachial (PB) area, located in the dorsolateral region of the pons (with the use of extracellular micropipette), in the anesthetized rat. Parabrachioamygdaloid (PA) neurons (n = 67) were antidromically identified after stimulation in the centralis nucleus of the amygdala (Ce). The axons of these neurons exhibit a very slow conduction velocity, between 0.26 and 1.1 m/s, i.e., in the unmyelinated range. 2. These PA neurons were located in a restricted region of the PB area: the subnuclei external lateral (PBel) and external medial (PBem). A relative somatotopic organization was found in this region. 3. These units were separated into two groups: 1) a group of nociceptive-specific (NS) neurons (69%), which responded exclusively to noxious stimuli, and 2) a group of nonresponsive (NR) neurons (31%). 4. The NS neurons exhibited low or lacked spontaneous activity. They responded exclusively to mechanical (pinch or squeeze) and/or thermal (waterbath or waterjet greater than 44 degrees C) noxious stimuli with a marked and sustained activation with a rapid onset and generally without afterdischarge. Noxious thermal stimuli generally induced a stronger response than the noxious mechanical stimuli. These neurons exhibited a clear capacity to encode thermal stimuli in the noxious range: 1) the stimulus-response function was always positive and monotonic; 2) the slope of the curve progressively increased up to a maximum where it was very steep, then the steepness of the slope decreased close to the maximum response; and 3) the mean threshold was 44.1 +/- 2 degrees C, and the point of steepest slope of the mean curve was around 47 degrees C. 5. The excitatory receptive fields of the NS neurons were large in the majority (70%) of the cases and included several areas of the body. A more marked activation was often obtained from stimuli applied to one part of the body, denoted as the preferential receptive field (PRF). In the other cases (30%), the excitatory receptive field was relatively small (SRF) and restricted to one part of the body (the tail, a paw, a hemiface, or the tongue). Both the PRF and SRF were more often located on the contralateral side. In addition, noxious stimuli applied outside the excitatory receptive field were found to strongly inhibit the responses of NS neurons. 6. All the NS neurons responded to intense transcutaneous electrical stimulation applied to the PRF or SRF with two peaks of activation.(ABSTRACT TRUNCATED AT 400 WORDS)
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The effect of systemic morphine on serotonin (5-HT) metabolism within the dorsal raphe nucleus (DRN) has been investigated by in vivo 5-hydroxyindole electrochemical (peak '3') detection in freely moving rats. Morphine caused a weak and delayed, but naloxone-reversible, increase in peak '3'. This increase was poorly, if at all, correlated with the morphine-induced analgesia. Finally, stress and/or noxious stimulation had no effect on this signal. These results are compared with our previous studies using the same methodological approaches and show that morphine caused a significant and specific increase in 5-HT metabolism at the levels of nucleus raphe magnus (NRM) and medullary dorsal horn. Furthermore, as shown in the present paper, there was also a good correlation between the time course of such increases and the analgesic effect of morphine. These findings are discussed with reference to the involvement of 5-HT mechanisms in the so-called DRN-NRM-dorsal horn 'intrinsic analgesic system'.
Several studies have considered the establishment of vascularization in intracerebral solid transplants of neural tissue. The widely supported interpretation of the results is that the vascular network of the solid grafts is already present before implantation into the host brain. The situation is different when dissociated fetal tissue is transplanted as a cell suspension because in these conditions the fetal vascular network is disrupted. The present study has, therefore, been undertaken to follow the angiogenesis in a transplant of dissociated fetal cells implanted into the excitotoxically neuron-depleted thalamus. The vascular network is compared to that observed in the intact and in the lesioned thalamus both in terms of morphology of the capillaries and of the function of the blood-brain barrier (BBB). In the transplant, capillaries, stained by Indian ink, are very few in number and have very fine calibers during the first 20 days after grafting. Some structures can be identified as immature blood vessels at the electron microscopic level. The blood vessels are progressively more numerous in the graft and they demonstrate mature ultrastructural features 2 months after grafting. Last, there is no leakage of the BBB for peroxidase. The vascularization seems to follow a pattern of maturation comparable to that described during development in the literature. In contrast, in the lesioned area, there is a reactive angiogenesis: 10 days after the excitotoxic injection (shortest time studied), there are many wide caliber vessels with expanded perivascular spaces engorged with mesodermal cells. A microvascularization also develops transiently during the first two months. Capillaries are abnormal from the functional point of view, since there is a leakage of the BBB to macromolecules. The use of an experimental model in which transplant had to grow in a lesioned area permits to determine two types of vascularization: an apparently normal developmental timetable, normal morphological and functional characteristics, in the transplant; a reactive angiogenesis, in the lesioned area.
Small iontophoretic application of wheat-germ-agglutinin conjugated to horseradish peroxidase, was used to simultaneously observe, in the rat, the afferent and the efferent connections of the cuneiformis nucleus (Cnf), in the same animal. The results demonstrate that the main Cnf afferent connections originate from: (1) 6 regions of the forebrain: the nucleus centralis of the amygdala, the zona incerta, the dorsomedial, ventromedial and the lateral area of the hypothalamus and the periventricular gray matter and (2) 4 regions of the midbrain: the substantia nigra pars lateralis, the peripeduncular area, the periaqueductal gray matter and the other side Cnf. In contrast to the wealth of origins of afferent fibers, most projections of the Cnf are concentrated to the B3 area and the bordering reticular nuclei. It is proposed that the Cnf, which mostly receives afferent connections from the forebrain and the midbrain, and which in turn densely projects on the B3 area, is a relay for the modulation of pain processes.
Small iontophoretic applications of wheat germ agglutinin-conjugated to horseradish peroxidase have been used to simultaneously observe, in the rat, the afferent and efferent connections of small parabrachial subregions in the same animal. The results demonstrate that a restricted region centered on the external portion of the lateral parabrachial area receives numerous afferents from the superficial laminae of the dorsal horn of the spinal cord and of the trigeminal subnucleus caudalis and sends a major projection to the nucleus centralis of the amygdala. This study suggests that the external portion of the lateral parabrachial area is a relay for messages evoked by noxious stimuli in a spino (trigemino)-ponto-amygdaloid pathway.
The aim of this electrophysiological investigation was to evaluate the activity of the spinal endogenous opioid systems in a chronic pain model, the arthritic rat. The activity of nociceptive non-specific dorsal horn neurons (n = 23) were recorded in 23 spinal unanesthetized decerebrated rats. Naloxone (1 mg/kg i.v.) induced a highly significant increase in the spontaneous firing rate of these neurons. This observation is in favor of a tonic activity of spinal opioid endogenous systems in such a disease. In addition, the same dose of naloxone facilitates the transmission of noxious messages at the spinal level as revealed by the large enhancement of the responses of these neurons to C-fiber stimulation. These results are in good agreement with behavioral data showing that such a relatively high dose of naloxone induced well-reproducible hyperalgesia and with some biochemical observations showing an increase of levels and biosynthesis of endogenous opioids in the spinal cord of the arthritic rat.
Highly selective tritiated ligands and autoradiography have been used to study mu and delta binding sites as well as neutral endopeptidase-24.11 in the dorsal horn of the rat spinal cord. Two weeks and 4 months after dorsal root rhizotomy, both opioid binding sites were dramatically decreased (congruent to 60%). In contrast, the level of neutral endopeptidase-24.11 remained unaltered.
The possible changes in neutral endopeptidase EC 3.4.24.11 ("enkephalinase", NEP), mu and delta opioid binding sites, were investigated using in vitro quantitative radioautography in various regions of the central nervous system of the Freund's adjuvant-induced arthritic rat, a model of chronic pain. Enkephalinase was labeled by a specific tritiated inhibitor, [3H]N-[(2RS)-3-hydroxyaminocarbonyl-2-benzyl-1-oxopropyl]glycine ([3H]HACBO-Gly), while mu and delta opioid binding sites were selectively labelled with [3H]Tyr-D-Ala-Gly-(Me)Phe-Gly-ol ([3H]DAGO) and [3H]Tyr-D-Thr-Gly-Phe-Leu-Thr ([3H]DTLFT), respectively. As compared to controls, no significant modifications were found in NEP, mu or delta binding sites at both supraspinal and spinal levels of arthritic rats. These results suggest that the enhanced efficiency of exogenous opioids or endogenous enkephalins, reported to occur in this model of chronic inflammatory pain, are not directly related to changes in mu and delta opioid binding sites or steady state levels of NEP.
In vitro superfusion of slices from the dorsal half of the rat cervical enlargement allowed the measurement of spontaneous, K+ (30 mM)- and capsaicin (0.5 microM)-evoked release of calcitonin gene-related peptide-like immunoreactive material (CGRPLI). The greater part of this immunoreactive material originated in primary afferent fibres since dorsal rhizotomy from C4 to Th2 (8 days before sacrifice) resulted in a 85-90% decrease in CGRPLI release. CGRPLI outflow which persisted after dorsal rhizotomy could still be enhanced by K+-induced depolarization but was no longer sensitive to the stimulatory effect of 0.5 microM capsaicin. Both delta (DTLET, D-Pen2-D-Pen5-enkephalin) and mu (DAGO, PL 017) opioid receptor agonists reduced the K+ evoked release of CGRPLI from the dorsal half of the cervical enlargement. Morphine was also inhibitory but the selective K opioid agonist U 69593 was inactive. As expected from the involvement of delta and mu receptors, the selective opioid antagonist ICI 174864 and naloxone prevented the inhibitory effects of DTLET and DAGO, respectively. These data suggest that opioid-induced presynaptic inhibiton of CGRP-containing primary afferent fibres may be involved in the analgesic effect of intrathecally injected delta and mu opioid agonists in rats.