Altered opioid-mediated control of the spinal release of dynorphin and met-enkephalin in polyarthritic rats.

Previous studies showed that spinal opioidergic neurotransmission is markedly altered in the polyarthritic rat, a model of chronic inflammatory pain. Present investigations aimed at assessing possible changes in opioid-mediated control of the spinal outflow of met-enkephalin (ME) and dynorphin (DYN) in these animals. Intrathecal (i.t.) perfusion under halothane anesthesia showed that polyarthritis was associated with both a 40% decrease in the spinal outflow of ME-like material (MELM) and a 90% increase in that of DYNLM. Local treatment with the mu-opioid agonist DAGO (10 microM i.t.) inhibited equally (-30%) the MELM outflow in polyarthritic and control rats, whereas the delta agonist DTLET (10 microM i.t.) also reduced the peptide outflow in controls (-27%) but enhanced it in polyarthritic animals (+56%). On the other hand, both DAGO (10 microM i.t.) and DTLET (10 microM i.t.) decreased (-40 and -49%) DYNLM outflow in polyarthritic rats, but were inactive in controls. Finally, neither MELM outflow nor that of DYNLM were affected by the kappa-agonist U50488H (10 microM i.t.) in both groups of rats. In all cases, the changes due to active agonists could be prevented by specific antagonists which were inactive on their own except the kappa antagonist nor-binaltorphimine (10 microM i.t.) that decreased (-38%) DYNLM outflow in polyarthritic rats. These data indicate that functional changes in spinal opioid receptors may promote enkephalinergic neurotransmission and reduce dynorphinergic neurotransmission in polyarthritic rats, thereby contributing to the analgesic efficacy of opioids in inflammatory pain.

Opposite changes in dopamine metabolites and met-enkephalin levels in the ventricular CSF of patients subjected to thalamic electrical stimulation.

High-frequency electrical stimulations of thalamic nuclei are currently used for the suppression of parkinsonian or essential tremor and for the relief of some types of intractable pain in man. However, the mechanisms by which such stimulations exert their therapeutic effects are essentially unknown. Attempts were made to provide some insight into these mechanisms by measuring the levels of the dopamine metabolites homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC), the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA) and met-enkephalin-like immunoreactivity in ventricular cerebrospinal fluid (CSF) of patients with Parkinson's disease (PD) or multiple sclerosis (MS) after a 30-minute therapeutic electrical stimulation of the ventralis intermedius nucleus of the thalamus. In nonstimulated control patients, the levels of these compounds did not significantly differ in two CSF samples taken 30 minutes apart. In stimulated patients, a decrease in dopamine metabolite levels associated with a relative increase in met-enkephalin-like immunoreactivity were observed in the CSF sample taken after the 30-minute stimulation as compared to the sample taken immediately before the stimulation. In contrast, the levels of 5-HIAA remained unaffected by the stimulation. These data confirmed the existence of negative interactions between dopaminergic and enkephalinergic systems in man similar to those previously demonstrated in rats. In addition, they suggest that alterations in dopaminergic or enkephalinergic neurotransmission might be involved in the therapeutic action of thalamic electrical stimulation in patients with parkinsonian symptoms and other patients.

Polyarthritis-associated changes in the opioid control of spinal CGRP release in the rat.

As a model of chronic inflammatory pain, Freund's adjuvant-induced polyarthritis has been shown to be associated with marked alterations in the activity of opioid- and calcitonin gene-related peptide (CGRP)-containing neurons in the dorsal horn of the spinal cord in rats. Possible changes in the interactions between these two peptidergic systems in chronic inflammatory pain were investigated by comparing the effects of various opioid receptor ligands on the spinal outflow of CGRP-like material (CGRPLM) in polyarthritic and age-paired control rats. Intrathecal perfusion of an artificial cerebrospinal fluid in halothane-anaesthetized animals allowed the collection of CGRPLM released from the spinal cord and the application of opioid receptor ligands. The blockade of kappa-opioid receptors similarly increased CGRPLM release in both groups of rats as expected of a kappa-mediated tonic inhibitory control of CGRP-containing fibres in control, as well as in polyarthritic rats. In contrast, the higher increase in CGRPLM outflow due to the preferential blockade of mu opioid receptors by naloxone in polyarthritic rats as compared to non-suffering animals supports the idea of a reinforced mu opioid receptor-mediated tonic inhibitory control of CGRP-containing fibres in rats suffering from chronic pain. Even more strikingly, the differences observed in the effects of delta-opioid receptor ligands on CGRPLM outflow suggest that delta receptors are functionally shifted from a participation in a phasic excitatory control in non-suffering rats to a tonic inhibitory control in polyarthritic rats. These data indicate that agonists acting at the three types of opioid receptors all exert a tonic inhibitory influence on CGRP-containing nociceptive primary afferent fibres within the spinal cord of polyarthritic rats. Such a convergence probably explains why morphine and other opioids are especially potent to reduce pain in subjects suffering from chronic inflammatory diseases.

Increased in vivo release of calcitonin gene-related peptide-like material from the spinal cord in arthritic rats.

Possible alterations in spinal systems containing calcitonin gene-related peptide (CGRP) due to polyarthritis were assessed in rats 3-4 weeks after an intradermal injection of Freund's adjuvant in the low back. The tissue levels of CGRP-like material (CGRPLM) were approximately 50% higher in the dorsal zone of the spinal cord and dorsal root ganglia at both the cervical and lumbar (but not thoracic) segments in polyarthritic rats than in age-paired control animals. In addition the rate of the spinal release of CGRPLM determined through an intrathecal perfusion procedure in halothane-anaesthetized animals was approximately 15-fold higher in polyarthritic rats than in controls. The blockade of mu-opioid receptors by intrathecal perfusion with 10 microM naloxone produced a larger increase in the spontaneous CGRPLM outflow in polyarthritic rats than in age-paired controls. Furthermore, the stimulation of mu-opioid receptors by intrathecal perfusion with 10 microM DAGO significantly inhibited the spinal outflow of CGRPLM only in polyarthritic rats. These data indicate that CGRP-containing primary afferent fibres are markedly activated in chronic suffering polyarthritic rats. This activation occurs in spite of an increased tonic inhibitory control by endogenous opioids acting at mu receptors.

Acupuncture-like stimulation induces a heterosegmental release of Met-enkephalin-like material in the rat spinal cord.

In order to investigate the effects induced by acupuncture on the activity of enkephalinergic neurons in the spinal cord, either the lumbar or the cervico-trigeminal area was perfused with artificial cerebrospinal fluid (CSF) (0.1 ml/min) in halothane-anaesthetized rats, and Met-enkephalin-like material (MELM) was measured in 0.5 ml fractions of the perfusates. The effects of manual acupuncture performed by a traditional Chinese acupuncturist at the 'Zusanli' point on the right hind limb were compared to the effects induced by acupuncture applied at a non-acupoint next to 'Zusanli.' The manipulation of needles either at the 'Zusanli' point or at the non-acupoint had no effect on the release of MELM from the lumbar area but significantly increased the release from the cervico-trigeminal zone. It is concluded that manual acupuncture triggers a heterosegmental activation of enkephalinergic neurones within the spinal cord and that this effect is non-specific in terms of the location of the stimulated point.

Neonatal capsaicin treatment abolishes the modulations by opioids of substance P release from rat spinal cord slices.

The possible modulation by opioids of substance P (SP) release at the spinal level was studied using slices of the dorsal half of the rat lumbar enlargement superfused with an artificial cerebrospinal fluid. Capsaicin (0.5 microM) selectively evoked a Ca2+-dependent overflow of SP-like material (SPLI) from primary afferent fibers which was enhanced in the presence of mu-opioid agonists (DAGO, FK 33824, sufentanyl, morphine), reduced by the delta-opioid agonist DTLET, and unaltered by the kappa-opioid agonist U 50488 H. Selective antagonists (naloxone, ICI 154129) prevented the effects of mu- and delta-opioid agonists. Neonatal capsaicin (50 mg/kg) abolished the stimulatory effect of in vitro capsaicin (0.5 microM) but not that of 30 mM K+ on SPLI outflow. This K+-induced SPLI release was unaffected by opioids. Presynaptic inhibitory control of SPLI release from capsaicin-sensitive primary afferent fibers might account for the analgesic effect of delta- but not mu- and kappa-opioid agonists at the spinal level.

Spontaneous and evoked release of met-enkephalin-like material from the spinal cord of arthritic rats in vivo.

Perfusion of the intrathecal space with artificial CSF was achieved in control and arthritic rats under halothane anaesthesia in order to collect the met-enkephalin-like material (MELM) released from the whole spinal cord. On the fourth week following the intradermal injection of Freund's adjuvant to induce arthritis, a marked reduction (-56%) in the spontaneous outflow of MELM was noted in arthritic rats. This effect did not involve changes in the degradation process of MELM, since it persisted when kelatorphan was added to the perfusing fluid in order to inhibit completely the peptidases acting on met-enkephalin. Raising the K+ concentration in the perfusing fluid from 2.4 to 40 mM, as well as moving the hind paws, produced a significant enhancement of MELM release which was (at least) as pronounced in arthritic as in control rats. These results suggest that the basal activity of spinal enkephalinergic neurones, but not that triggered by various stimuli, is reduced in arthritic rats.

Opioid receptors and neuropeptides in the CNS in rats treated chronically with amoxapine or amitriptyline.

The central mechanism responsible for the potentiation by antidepressant drugs of analgesia induced by morphine, was explored by measuring the levels of various neuropeptides (met-enkephalin, leu-enkephalin, dynorphin, substance P and cholecystokinin-like materials) and the density of delta and mu opioid binding sites in the spinal cord of rats treated for 14 days with amoxapine (10 mg/kg i.p., daily) or amitriptyline (10 mg/kg i.p., daily). Similar measurements were made in the hypothalamus and cerebral cortex for comparison. Chronic treatment with amoxapine or amitriptyline did not affect the levels of dynorphin, substance P and cholecystokinin, but markedly enhanced the levels of leu-enkephalin in the three structures examined. The levels of met-enkephalin were also increased after treatment with amitriptyline but only in the spinal cord and hypothalamus. No changes in opioid receptors were found in the cerebral cortex, but the densities of delta and mu opioid binding sites were increased in the spinal cord, and decreased in the hypothalamus of rats treated with amoxapine or amitriptyline. These changes induced by antidepressants in opioidergic markers at the spinal level might account for the potentiation of the action of morphine in amoxapine- or amitriptyline-treated rats. In addition, the observed alterations in the same markers in the hypothalamus could be associated with changes induced by antidepressants in neuroendocrine regulation.

Opposite effects of delta and mu opioid receptor agonists on the in vitro release of substance P-like material from the rat spinal cord.

Superfusion of slices from the dorsal half of the lumbar enlargement of rat spinal cord with Krebs-Henseleit medium supplemented with 30 microM bacitracin allowed the collection of substance P-like immunoreactive material (SPLI), which was released at a rate of approximately 10 pg/4 min. Tissue depolarization by an excess of K+ (30-60 mM) or veratridine (50 microM) induced a marked increase in SPLI outflow, provided that Ca2+ was present in the superfusing fluid. K+- or veratridine-induced SPLI overflow could be modulated in opposite directions by mu and delta opioid receptor agonists. Thus, the two preferential mu agonists Tyr-D-Ala-Gly-MePhe-Gly-ol (DAGO; 10 microM) and Tyr-D-Ala-Gly-MePhe-Met(O)5-OH (FK-33824; 0.1 microM) enhanced SPLI overflow from depolarized tissues, whereas the selective delta agonists Tyr-D-Thr-Gly-Phe-Leu-Thr (deltakephalin; 3 microM) and [2-D-penicillamine, 5-D-penicillamine]enkephalin (50 microM) reduced it. The effect of DAGO was antagonized by a low concentration (1 microM) of naloxone but not by the selective delta antagonist ICI-154129 (50 microM). In contrast, the latter drug prevented the inhibitory influence of delta agonists on K+-induced SPLI release. Complementary experiments with morphine (10 microM) and [2-D-alanine, 5-D-leucine]enkephalinamide (3 microM), in combination with 1 microM naloxone or 50 microM ICI-154129 for the selective blockade of mu or delta receptors, respectively, confirmed that the stimulation of mu receptors increased, whereas the stimulation of delta receptors reduced, SPLI overflow. The results suggest that, at the spinal level, and antinociceptive action of delta but not mu agonists might involve a presynaptic inhibition of substance P-containing primary afferent fibers.

Local and remote effects of intra-caudate administration of GABA-related drugs on Met-enkephalin release in the basal ganglia.

The possible influence of GABAergic systems on the activity of enkephalinergic neurones within the basal ganglia was examined by measuring the release of Met-enkephalin in the caudate nuclei and pallida of halothane-anesthesized cats treated by intra-caudate applications of GABA-related drugs. Depending on the concentration used, GABA exerted local stimulatory (at 10 microM of the amino acid) or inhibitory (at 0.5 mM) action on Met-enkephalin release in the cat caudate nucleus. Only the inhibition was reproduced by the GABA agonists muscimol (1 microM) and (-)-balcofen(10 microM) and by diazepam 10 microM). Conversely, the intra-caudate application of the GABA antagonist bicuculline enhanced markedly the local release of the pentapeptide. Complementary studies using slices of the rat striatum (caudate nucleus + putamen) revealed that a low concentration of GABA (10 microM) tended to increase the K+-evoked efflux of Met-enkephalin, whereas a high concentration of the amino acid exerted a strong inhibitory effect on the peptide release. Such in vivo and in vitro findings suggest that the GABA-induced inhibition of Met-enkephalin release took place via the stimulation of specific GABA A and GABA B receptors within the caudate nucleus, whereas the GABA-induced increase of the peptide release might involve some intracellular regulatory processes in striatal neurones containing both GABA and enkephalins. In addition to altering the local release of Met-enkephalin, intra-caudate applications of GABA-related drugs affected the peptide release in the ipsilateral globus pallidus and contralateral basal ganglia. The observed changes suggest that GABA A, but not GABA B, receptors participated in some tonic inhibitory influence of striatal GABAergic neurones on the striato-pallidal enkephalinergic system. Furthermore, the present results confirmed previous studies (Bourgoin et al.) showing that GABAergic neurones can contribute to some bilateral modulation of enkephalinergic neurones within the basal ganglia.

Biochemistry of the hypothalamus in Parkinson's disease.

We assayed the content of neurotransmitters (or their synthesizing enzymes) and neuropeptides in the hypothalamus of control and parkinsonian brains post mortem. Only dopamine concentrations were lower than normal in Parkinson's disease, suggesting that deficiency in hypothalamic dopamine transmission may play a role in the autonomic and endocrine abnormalities of this disorder.

[Determination of pituitary hormones in cerebrospinal fluid in normal subjects and in hypothalamo-pituitary diseases. Secreting or non-secreting pituitary adenoma, empty sella syndrome, hypothalamic syndromes]. / Le dosage des stimulines hypophysaires dans le liquide céphalo-rachidien chez le sujet normal et en pathologie hypothalamo-hypophysaire. Adénome hypophysaire sécrétant ou non sécrétant, selle turcique vide, syndromes hypothalamiques.

Cerebrospinal fluid (CSF) and serum concentrations of TSH, ACTH, FSH, LH, GH and PRL were measured simultaneously in 34 subjects divided into 3 groups: I--12 normal subjects (6 males and 6 females); II--12 prolactin adenomas (3 males and 9 females); III--5 empty sella syndromes; 3 hypothalamic disorders; 1 chromophobe adenoma; 1 pituitary dwarfism. It is concluded that:--pituitary hormones are the normal constituents of CSF but the level can be undetectable and in any case lower than the serum level; --there is a positive correlation between serum and CSF concentration of PRL when serum PRL is higher than 20 ng/ml, indicating that the CSF level is influenced by serum level; --in prolactin adenomas only prolactin is elevated in the CSF; --there is no correlation between the high level of CSF-PRL and a suprasellar extension of the adenoma.