Dynorphin Content Research Articles

The role of opioids in newborn pig fluid percussion brain injury

The present study was designed to characterize the relationship between cerebral opioid concentration, cerebral hemodynamics, and cerebral oxygenation following percussion brain injury in neonatal pigs. Previous research found that opioids represent a significant vasoactive component in the regulation of the neonatal piglet cerebral circulation. Anesthesized newborn (1–5 days old) pigs equipped with a closed cranial window were connected to a percussion device consisting of a saline-filled cylindrical reservoir with a metal pendulum. Brain injury of moderate severity (1.9–2.3 atm.) was produced by allowing the pendulum to strike a piston on the cylinder. Fluid percussion brain injury decreased pial arteriolar diameter (132 ± 5 to 110 ± 5 μm within 10 min). Cerebral blood flow also fell within 10 min of injury and continued to fall progressively for 3 h, resulting in a 46 ± 4% decrease. Within 30 s of brain injury, there was a transient increase in cerebral hemoglobin-O2 saturation that was reversed to a progressive profound decrease in cerebral hemoglobin-O2 saturation for the next 3 h, as measured by near infrared spectroscopy. CSF opioid concentrations were increased 10 min after brain injury; dynorphin showed the largest proportional increase (5.8 ± 0.9 fold). The CSF concentration for other opioids continued to increase over 180 min while the dynorphin concentration progressively decreased with time. In naloxone (1 mg/kg i.v.) pretreated piglets, the brain injury induced decrease in arteriolar diameter was attenuated (129 ± 5 to 121 ± 5 μm within 10 min). Similarly, the decrease in regional cerebral blood flow and cerebral hemoglobin-O2 saturation observed following brain injury were also blunted by naloxone. These data show that CSF opioid concentrations increase following brain injury and that the time course and relative increase in CSF concentration vary from opioid to opioid. These data also indicate that in the immature animal, opioids contribute to arteriolar constriction, decreased cerebral blood flow, decreased cerebral oxygenation, and could play a role in causing ischemia after brain injury.

Decreased concentration of dynorphin a in striatum and increased activity of dynorphin converting enzyme in the spinal cord following repeated morphine treatment

This study was designed to explore the possible correlation between the activity of dynorphin A converting enzyme (DCE), substance P converting endopeptidase (SPE) or the concentration of opioid peptides and substance P (1-11) in the mouse striatum, hippocampus and spinal cord with the intensity of opioid dependence following either single (short-term dependence) or threefold (long-term dependence) implantations of morphine pellets. It was found that the DCE activity was increased in the lumbar part of spinal cord during withdrawal after single, but not threefold morphine pellet implantations. The concentration of dynorphin A in the striatum was decreased during abstinence after threefold pellet implantations

Stress, arousal and opioid peptide-like immunoreactivity in restricted- and ad lib.-fed broiler breeder fowls

1. 1. From 2 weeks of age, female broiler breeders were fed on one of three treatments: a commercial restricted ration (R), twice that amount (2R), or ad lib. (AL). With R and 2R, birds were fed daily at 9 a.m. and ate all their food in <15 min. 2. 2. Blood samples were taken every fourth week from 5 to 21 weeks. Two putative indices of stress, basophil frequency and the heterophil/lymphocyte ratio, were highest with R and lowest with AL, consistently so with the former index and in all but the last week with the latter. Another stress index, plasma corticosterone concentration, was not affected by feeding treatment. 3. 3. Tonic immobility duration, an index of fear, was lower with R and 2R than AL at 7 and 12 weeks, but not 21 weeks. Headflick frequency, an index of arousal, was lower with R than 2R and AL, during 3 min silence and 3 min noise, at 11, 15 and 21 weeks. 4. 4. At 21 weeks, intramuscular injection of nalmefene, an antagonist of central opioid receptors, caused increases in headflick frequency, but there was no significant interaction with feeding treatment in this response. 5. 5. Feeding treatment had no significant effect on levels of β-endorphin- or leu-enkephalin-like immunoreactivity in brains of birds killed at 21 weeks, but dynorphin concentration was higher with 2R. Opioid receptor density, which could have been affected, was not measured. 6. 6. It is concluded that the commercial feeding programme caused physiological evidence of stress, and apparent reductions in fearfulness and arousal. Results of the tests at 21 weeks did not rule out the possibility that the reduced arousal was associated with altered opioid status.

Comparative effects on blood pressure and heart rate of dynorphin A(1–13) in anterior hypothalamic area, posterior hypothalamic area, nucleus tractus solitarius, and lateral cerebral ventricle in the rat

The objective of this study was to explore the effects of the endogenous opioid peptide dynorphin A(1–13) on the CNS regulation of blood pressure and heart rate. Wistar rats, anesthetized with pentobarbital and halothane, received dynorphin A(1–13) microinjected into the anterior hypothalamus area (AHA), the posterior hypothalamic area (PHA), the nucleus tractus solitarius (NTS), or the lateral cerebral ventricle (ICV). Dynorphin A(1–13), 20 (12 nmol) or 30 μg ICV, produced significant ( p < 0.05) reductions in blood pressure and heart rate. Naloxone, 50 μg/kg ICV, completely prevented the blood pressure response and significantly ( p < 0.05) blunted the heart rate response to the highest dynorphin concentration, 30 μg ICV (18 nmol). Dynorphin A(1–13), 5 μg, in the NTS significantly ( p < 0.05) decreased systolic and diastolic blood pressure and heart rate with the response being evident 10 min and persisting for 30 min after injection. In contrast, the same dose of dynorphin A(1–13) in the AHA produced an immediate, marked, and significant ( p < 0.05) decrease in systolic and diastolic blood pressure and heart rate that attained its maximum 1–3 min and returned rapidly towards baseline levels. Dynorphin A(1–13), 5 or 10 μg in the posterior hypothalamic area, was not associated with any change in blood pressure or heart rate. Injection of the diluent at any site was not associated with any changes in blood pressure or heart rate. The maximum change in blood pressure with dynorphin was greater in the AHA than NTS, and the maximum change in heart rate was greater in the NTS than AHA. These data indicate a potential role for dynorphin as a modulator of the CNS regulation of blood pressure and cardiac rate, and this is mediated in part through different areas in the brain that maybe localized to the anterior hypothalamic area and nucleus tractus solitarius but not the posterior hypothalamic area.

Spinal cord dynorphin: positive region-specific modulation during pregnancy and parturition

In laboratory animals and humans, pregnancy is associated with opioid-mediated elevations in the threshold for responsiveness to aversive stimuli. Previous pharmacological analysis has demonstrated that this analgesia results, at least in part, from the activation of spinal cord kappa opioid receptors utilizing dynorphin as the major opioid substrate. The present report demonstrates that during late pregnancy, the content of spinal dynorphin A(1–17 and 1–8) is altered in a region-specific fashion. As a result, levels of dynorphin peptides are elevated, but only in the lumbar spinal region. In parturient animals, lumbar levels of dynorphin A(1–8) remained elevated but there was an additional increment in the content of dynorphin A(1–17). During late gestation, spinal content of Met-enkephalin and its precursor are also elevated, but, in contrast to dynorphin peptides, there is no interaction between condition and spinal level. Possible analgesic synergy between μ−δ and κ opioid receptor systems is discussed. It is concluded that some parameter(s) of the pregnant condition triggers the activation of a spinal cord dynorphin system that attenuates the pain associated with late pregnancy and labor.

CSF neuropeptides in cancer pain: effects of spinal opioid therapy.

The cerebrospinal fluid (CSF) levels of the opioid peptides met-enkephalin (ME), beta-endorphin (BE) and dynorphin (DYN) as well as the putative sensory neuropeptides substance P (SP), somatostatin (SOM), calcitonin gene related peptide (CGRP) and vasoactive intestinal polypeptide (VIP) were determined in 10 patients with severe nociceptive pain due to malignancy, before and after initiation of spinal opioid therapy, and in 10 control patients. Pain intensity, evaluated by means of a 100-mm visual analog scale (VAS), was reduced from 39 +/- 9 to 18 +/- 10 for continuous pain and from 70 +/- 10 to 10 +/- 8 for intermittent pain (means +/- s.e.mean). Lumbar CSF immunoreactive ME and DYN concentrations were significantly increased (P = 0.05) and BE and VIP were significantly decreased (P < or = 0.05) in the pain patients. A slight, but non-significant (P = 0.06) decrease in SP-like immunoreactivity was found after initiation of spinal opioid therapy. Visceral pain seemed to be associated with low immunoreactive SP and ME concentrations compared to somatic pain. A highly significant correlation was found between SP and ME (P < 0.001) and to a lesser extent also between other peptides. We conclude that the concentrations of the endogenous opioids were more affected by nociceptive pain states than the non-opioid peptides. The origin of pain may also influence the results. The postulated inhibition of peptide release by spinal opioid application seemed to be present for SP, but could otherwise not be confirmed.

Endogenous opioid regulation of oxytocin secretion through pregnancy in the rat.

We have investigated the influence of endogenous opioids on oxytocin secretion during pregnancy. In blood-sampled conscious rats on days 18 and 21 of pregnancy plasma oxytocin concentration, measured by radioimmunoassay, was significantly increased compared to non-pregnant or post-partum rats. On days 15, 18 and 21 of pregnancy but not in non-pregnant, early pregnant or post-partum rats, the opioid antagonist naloxone caused a significant increase in plasma oxytocin compared to vehicle injection, indicating activation of an endogenous opioid restraint over oxytocin secretion. Electrically stimulated neural lobes isolated from 16- and 21-day pregnant rats released more oxytocin than those from non-pregnant rats. However, naloxone (10(-5) M) was less effective at potentiating, and the kappa-opioid agonist U50,488 (10(-5)M) was less effective at inhibiting, stimulated release at the end of pregnancy than in non-pregnant rats suggesting desensitization of oxytocin nerve terminals to actions of endogenous opioids. Neural lobes from male rats drinking 2% saline for 4 days also showed desensitization of oxytocin nerve endings to naloxone. Neither neural lobe content of dynorphin A(1-8), an endogenous kappa-opioid, nor prodynorphin mRNA expression, measured by in situ hybridization histochemistry in the supraoptic nucleus altered during pregnancy. However, neural lobe content of Met5-enkephalin significantly decreased by day 21 of gestation suggesting enhanced release.(ABSTRACT TRUNCATED AT 250 WORDS)

17 beta-estradiol and progesterone positively modulate spinal cord dynorphin: relevance to the analgesia of pregnancy.

Pregnancy and parturition are associated with opioid-mediated elevations in maternal pain thresholds. This analgesia is subserved by a spinal cord dynorphin/kappa-opiate receptor system. During gestation, elevated pain thresholds are paralleled by a significant increase in the content of dynorphin (1-17 and 1-8) in the lumbar spinal cord. An additional increment in lumbar dynorphin (1-17) concentration, but not that of dynorphin (1-8), occurs in parturient animals. Simulation of the pregnancy blood concentration profile of 17 beta-estradiol and progesterone ('hormone-simulated pregnancy') also results in an opioid analgesia, the magnitude and temporal pattern of which closely approximates that of actual gestation. The current study demonstrates that during hormone-simulated pregnancy, the spinal cord content of dynorphin (1-17) [but not dynorphin (1-8)] is positively modulated. This regulation is time- (or dose-)-dependent and region-specific. Significant elevations in spinal levels of dynorphin (1-17) are observed during steroid dose periods 3 and 4, corresponding to the last week of actual gestation and parturition, respectively. Increased dynorphin (1-17) content is observed in only the lumbar spinal region. These changes are temporally and anatomically identical to those which occur during actual gestation and parturition. It is concluded that changes in circulating 17 beta-estradiol and progesterone, that occur as a natural consequence of gestation, activate a dynorphin system in the lumbar spinal cord. This attenuates the pain associated with late pregnancy and labor. The pattern of circulating sex steroids can be an important determinant of the activity of central opioid analgesic systems.

Chronic selective activation of excitatory opioid receptor functions in sensory neurons results in opioid ‘dependence’ without tolerance

We previously showed that mouse sensory dorsal root ganglion (DRG) neurons chronically exposed to 1 μM d-ala 2- d-leu 5-enkephalin (DADLE) or morphine for > 2–3 days in culture become tolerant to the usual opioid inhibitory receptor-mediated effects, i.e. shortening of the duration of the calcium-dependent component of the action potential (APD), and supersensitive to opioid excitatory APD-prolonging effects elicited by low opioid concentrations. Whereas nanomolar concentrations of dynorphin(1–13) or morphine are generally required to prolong the APD of naive DRG neurons (by activating excitatory opioid receptors), femtomolar levels become effective after chronic opioid treatment. Wheareas 1–30 nM naloxone or diprenorphine prevent both excitatory and inhibitory opioid effects but do not alter the APD of naive DRG neurons, both opioid antagonists unexpectedly prolong the APD of most of the chronic opioid-treated cells. In the present study, chronic exposure of DRG neurons to 1 μM DADLE together with cholera toxin-B subunit (which selectively blocks GM1 ganglioside opioid excitatory, but not inhibitory, receptor functions) prevented the development of opioid excitatory supersensitivity and markedly attenuated tolerance to opioid inhibitory effects. Conversely, sustained exposure of DRG neurons to 1 nM DADLE, which selectively activates excitatory opioid receptor functions, resulted in characteristic opioid excitatory supersensitivity but no tolerance. These results that ‘dependence’-like properties can be induced in chronic opioid-treated sensory neurons in the absence of tolerance. On the other hand, development of some components of tolerance in these cells may require sustained activation of both excitatory, as well as inhibitory, opioid receptor functions.

Dynorphin A content in the rat brain and spinal cord after a localized trauma to the spinal cord and its modification with p-chlorophenylalanine : An experimental study using radioimmunoassay technique

The distribution of dynorphin A in the spinal cord and brain of normal rats and of rats subjected to a focal injury of the spinal cord was examined in a rat model using a radioimmunoassay (RIA) technique. The validity of RIA was checked by high performance liquid chromatography (HPLC). Furthermore, the possibility that the peptide is somehow functionally related with endogenous 5-hydroxytryptamine (5-HT, serotonin), was also evaluated using a pharmacological approach. In normal animals, the peptide content was very similar in the spinal cord segments (T 9, T 10–11, and T 12) examined whereas, the dynorphin content of the whole brain was about two-fold higher compared with that in the spinal cord. A focal injury to the spinal cord in the right dorsal horn (about 1.5 mm deep, 2.5 mm long and 1.5 mm to the right of the midline) of the lower thoracic cord (T 10–11) in urethane anaesthetised animals significantly altered the peptide content in the whole brain as well as in the spinal cord. Thus, a decrease in the peptide level in whole brain, T 10–11 and in the T 12 segments of the spinal cord was observed 1 and 2 h after trauma. At 5 h, the peptide had accumulated markedly in the T 9 segment (about a two-fold increase) as compared with the controls. At this time, the peptide content had been restored in the T 10–11 and T 12segments. On the other hand, the whole brain dynorphin level continued to remain low (by 55%) as compared to the control group. Pretreatment with p-chlorophenylalanine (p-CPA, a 5-HT synthesis inhibitor) prevented the increase of the peptide in the T 9 segment of the cord seen after 5 h injury. The peptide level of the other two spinal cord samples and the whole brain was similar to that of untreated traumatised rats. Furthermore, p-CPA treatment alone (in untraumatised animals) significantly reduced the whole brain dynorphin content (45% decrease) without having any effect on the spinal cord peptide level. Our results, for the first time, provide biochemical evidence that (i) dynorphin actively participates in the tissue response to a spinal cord trauma, (ii) its concentration is widely altered in the spinal cord and brain after a focal trauma to the cord, and (iii) suggest that a functional interaction between serotonin and dynorphin exists in the central nervous system.

Dopaminergic modulation of striatal neuropeptides: differential effects of D 1 and D 2 receptor stimulation on somatostatin, neuropeptide Y, neurotensin, dynorphin and enkephalin

Dopaminergic modulation of neuropeptides in rat striatum was investigated by examining the effects of prolonged D 1 or D 2 receptor stimulation on levels of somatostatin, neuropeptide Y, neurotensin, dynorphin and enkephalin. Rats with a unilateral 6-hydroxydopamine (6-OHDA) lesion of the nigrostriatal pathway were treated for 7 days with either the D 1 agonist SKF 38393 (12.5 mg/kg/day) or the D 2 agonist quinpirole (1 mg/kg/day). Two regimens of agonist treatment were compared: continuous infusion via osmotic pump implanted i.p. and intermittent (once daily) i.p. injection. Rats were sacrificed 3 h after the last injection and peptide levels measured in the striatum bilaterally by radioimmunoassay; alterations in peptide content were observed primarily in the denervated striatum. In comparison to values from lesioned, vehicle-treated controls, intermittent administration of SKF 38393 reduced somatostatin Y (down 61% and 57%, respectively), increased neurotensin (up 105%) and dynorphin (up 184%) and had no effect on enkephalin; continuous SKF 38393 decreased neuropeptide Y by 39% but did not alter levels of the other peptides. Continuous quinpirole elevated somatostatin and neuropeptide Y levels (up 43% and 33%, respectively), but reduced the lesion-induced increases in both neurotensin (down 51%) and enkephalin (down 24%) content. Conversely, intermittent quinpirole decreased somatostatin (down 35%) and neuropeptide Y (down 27%), increased neurotensin content by 79% and had no effect on enkephalin. Dynorphin levels were not altered by either continuous or intermittent quinpirole. These findings reveal the complexity of dopaminergic influences on striatal neuropeptides. Somastostatin, neuropeptide Y and neurotensin levels are modulated by both D 1 and D 2 dopamine receptors: somatostatin and neuropeptide Y show parallel changes, but the alterations in neurotensin are opposite in direction to those of somatostatin and neuropeptide Y. Striatal dynorphin content is influenced only by D 1 receptor stimulation, while enkephalin is modulated only by D 2 receptors. The data also demonstrate that striatal neuropeptides are profoundly influenced by the temporal characteristics of dopamine receptor stimulation.

After chronic opioid exposure sensory neurons become supersensitive to the excitatory effects of opioid agonists and antagonists as occurs after acute elevation of GM1 ganglioside

Mouse sensory dorsal-root ganglion (DRG) neurons chronically exposed to 1 μM D-Ala 2-D-Leu 5-enkephalin (DADLE) for >1 week in culture become tolerant to opioid inhibitory effects, i.e. shortening of the duration of the calcium-dependent component of the action potential (APD). Acute application of higher concentrations of DADLE (ca. 10 μM) to these treated neurons not only fails to shorten the APD but, instead, generally elicits excitatory effects, i.e. prolongation of the APD. The present study shows that chronic DADLE- or morphine-treated DRG neurons also become supersensitive to the excitatory effects of opioids. Whereas nM concentrations of dynorphin(1–13) are generally required to prolong the APD of naive DRG neurons, fM levels become effective after chronic opioid treatment. Whereas 1–30 nM naloxone or diprenorphine do not alter the APD of naive DRG neurons, both opioid antagonists unexpectedly prolong the APD of most of the treated cells. Similar supersensitivity to the excitatory effects of opioid agonists and antagonists was previously observed after acute treatment of naive DRG neurons with GM1 ganglioside. Our results suggest that both chronic opioid and acute GM1 treatments of DRG neurons greatly enhance the efficacy of opioid excitatory receptor functions so that even the extremely weak agonist properties of naloxone and diprenorphine become effective in prolonging the APD of these treated cells when tested at low concentrations, whereas their antagonist properties at inhibitory opioid receptors do not appear to be altered. Furthermore, whereas cholera toxin-B subunit (CTX-B; 1–10 nM) blocks opioid-induced APD prolongation in naive DRG neurons (presumably by interfering with endogenous GM1 modulation of excitatory opioid receptors functions), even much higher concentrations of CTX-B were ineffective in chronic opioid-treated as well as acute GM1-elevated neurons. These and related data suggest that opioid excitatory supersensitivity in chronic opioid-treated DRG neurons may be due to a cyclic AMP-dependent increase in GM1 ganglioside levels. Our results may clarify mechanisms of opioid dependence and the paradoxical supersensitivity to naloxone which triggers withdrawal symptoms after opiate addiction.

Stress-induced changes in brain Met-enkephalin, Leu-enkephalin and dynorphin concentrations

Methionine-enkephalin (Met-enkephalin), leucine-enkephalin (Leu-enkephalin) and dynorphin A (1–17) (dynorphin A) concentrations in discrete brain areas were determined in the mice showing behavioral changes induced by stress using radioimmunoassay (RIA). In the present experiment, we used environment-induced conditioned suppression of motility and forced swimming-induced immobility. In the environment-induced conditioned suppression of motility, Met-enkephalin concentration in the striatum and hypothalamus significantly decreased. Leu-enkephalin concentration in the hypothalamus also decreased. Dynorphin A concentraion in the striatum decreased, but significantly increased in the hypothalamus and pituitary. In the forced swimming-induced immobility, Met-enkephalin concentration in the striatum significantly decreased. Leu-enkephalin concentration in the hypothalamus and pituitary significantly decreased. Dynorphin A concentration in the pituitary decreased, but significantly increased in the hypothalamus. Our results indicated that the concentrations of Met-enkephalin, Leu-enkephalin and dynorphin A in the discrete brain areas changed in two different stressful situations. These findings suggested that these peptides might modulate the behavioral changes induced by stressors.

Decreased glutamate release correlates with elevated dynorphin content in the hippocampus of aged rats with spatial learning deficits

The effects of aging on extracellular glutamate and tissue dynorphin content in the hippocampus were examined in Fischer-344 rats. Young adult (4-month-old) and aged (24-month-old) rats were trained to find a hidden platform in the Morris water task. Aged rats were unable to acquire the spatial learning task as rapidly as young controls. Following behavioral testing, an in vivo microdialysis perfusion method was used to determine extracellular glutamate levels in the hippocampus. There was a 25-35% reduction in extracellular glutamate concentration in both dorsal and ventral hippocampus of aged rats compared to young rats, in the absence of any change in tissue glutamate levels. Radioimmunoassay showed an increase in dynorphin A(1-8)-like immunoreactivity [DYN-A(1-8)LI] in both dorsal and ventral hippocampus, but not striatum, of aged rats. Immunocytochemistry indicated that this increase was localized to the dentate granule cells and mossy fibers. Furthermore, among the aged rats the increase in DYN-A(1-8)LI was inversely correlated with the decrease in extracellular glutamate. These results suggest that the disregulation of dynorphin observed in cognitively impaired aged rats is related to reduced excitatory transmission within the hippocampal formation.

Increased β-endorphin and dynorphin concentrations in discrete hypothalamic regions of genetically obese (ob/ob) mice

Disturbances in hypothalamic β-endorphin and dynorphin levels were investigated in non-fasted genetically obese ( ob/ob) and homozygous lean mice at 14–15 weeks of age. Eight brain regions were microdissected from fresh, unfixed brain slices, and opioid peptide concentrations were determined in tissue micropunches by radioimmunoassay. A two-fold increase in β-endorphin levels in ob/ob versus lean mice were found in the ventromedial and dorsomedial hypothalamic nuclei respectively. Dynorphin levels were comparable between ob/ob and lean mice in the anterior, lateral, ventromedial and paraventricular hypothalamic areas, but a 5-fold increase in dynorphin concentrations was detected in the dorsomedial hypothalamic nucleus of the ob/ob mouse. These results demonstrate that increased concentrations of β-endorphin and dynorphin occur in discrete hypothalamic nuclei, which are known to influence food intake and glucose homeostasis. This could signify an important central defect contributing to hyperphagia and glucoregulatory dysfunction in obese mice.

Levodopa replacement therapy alters enzyme activities in striatum and neuropeptide content in striatal output regions of 6-hydroxydopamine lesioned rats

The effects of striatal dopamine denervation and levodopa replacement therapy on neuronal populations in the rat striatum were assessed by measurement of glutamic acid decarboxylase (GAD) and choline acetyltransferase (CAT) activities in the striatum, dynorphin and substance P concentrations in the substantia nigra, and enkephalin concentration in the globus pallidus. Rats with a unilateral 6-hydroxydopamine (6-OHDA) lesion of the nigrostriatal pathway were treated for 21 days with levodopa (100 mg/kg/day, i.p., with 25 mg/kg benserazide) on either an intermittent (b.i.d.) or continuous (osmotic pump infusion) regimen and sacrificed following a three day drug washout. In saline-treated control rats, striatal GAD activity and globus pallidus enkephalin content were elevated and nigral substance P content was reduced ipsilateral to the 6-OHDA lesion. Intermittent levodopa treatment further increased GAD activity, decreased CAT activity, restored substance P to control levels, markedly increased dynorphin content, and had no effect on enkephalin. In contrast, continuous levodopa elevated globus pallidus enkephalin beyond the levels occurring with denervation had no effect on any of the other neurochemical measures. These results indicate that striatal neuronal populations are differentially affected by chronic levodopa therapy and by the continuous or intermittent nature of the treatment regimen. With the exception of substance P, levodopa did not reverse the effects of the 6-OHDA lesion but, rather, either exacerbated the lesion-induced changes (e.g. GAD and enkephalin) or altered neurochemical markers which had been unaffected by the lesion (e.g. CAT and dynorphin). These findings suggest that chronic levodopa treatment may create a new functional state in the striatum which is different from both the normal state and the denervated state and may provide insight into the pathophysiology of the motor response complications which often accompany long-term levodopa therapy in parkinsonian patients.

Localisation and pharmacological characterisation of [ 3H]bremazocine binding in the bovine adrenal medulla

[ 3H]Bremazocine (5 nM), in the presence of excess unlabelled μ and δ opioid ligands labelled two anatomically distinct populations of binding sites in the bovine adrenal medulla; a high density over the peripheral adrenaline-containing region of the medulla and a lower density over the central noradrenaline-containing region. This non-μ, non-δ opioid binding was specific (diprenorphine sensitive) but did not appear to involve classical κ ( κ 1), σ or PCP binding sites being insensitive to high concentrations of dynorphin (1–13), 3-PPP or MK-801. A significant proportion of the binding at both locations was however sensitive to competition by U50,488H or metorphamide. These data provide further evidence to support the existence of multiple opioid binding sites in the bovine adrenal medulla.

Nigrostriatal dopamine mediates the stimulatory effects of corticotropin releasing factor on methionine-enkephalin and dynorphin release from the rat neostriatum

This study examined the effects of corticotropin-releasing factor (CRF) on the in vitro release of methionine-enkephalin (Met-enkephalin) and dynorphin in neostriatal slices taken from rats with unilateral 6-hydroxydopamine (6-OHDA)-induced lesions of the nigrostriatal DA pathway. In neostriatal slices from control saline-infused animals and in those from the contralateral hemisphere of 6-OHDA-lesioned animals, CRF (10 −10 M) administered as a 90-min pulse exerted potent stimulatory effects on both Met-enkephalin and dynorphin release. In the neostriatal slices of 6-OHDA-lesioned striata, both the basal release and tissue content of Met-enkephalin were significantly ( P < 0.01) higher (2-fold) than those of control animals and the contralateral hemisphere of 6-OHDA-lesioned animals; however, neither the basal release nor the tissue content of dynorphin in 6-OHDA-lesioned striata was significantly different from control striata. In response to CRF (10 −10 M) the release of both Met-enkephalin and dynorphin were significantly diminished in slices of 6-OHDA-lesioned striata. These data support previous studies suggesting that nigrostriatal DA itself may exert a tonic inhibitory action on the activity of striatal Met-enkephalin neurones; however, DA may not have the same influence on striatal dynorphin neurons. However, the results of this study demonstrate that the action of CRF on Met-enkephalin as well as dynorphin release from the rat neostriatum is DA dependent. The data suggest that CRF receptors in the rat neostriatum may be localized on nigrostriatal/nigropallidal DA terminals/collaterals.

Dynorphin prolongs the action potential of mouse sensory ganglion neurons by decreasing a potassium conductance whereas another specific kappa opioid does so by increasing a calcium conductance

Previous studies have reported that large (μM) concentrations of kappa opioids, e.g. dynorphin and 3,4 dichloro- N-methyl- N-(-2[1-pyrrolidinyl]-cyclohexyl)benzene-acetamide (U-50.488H), shorten the duration of the calcium component of the action potential of dorsal root ganglion neurons by decreasing a voltage-sensitive Cs 2+ conductance. The present study showed that, in addition to these inhibitory modulatory effects, small (nM) concentrations of dynorphin, as well as U-50.488H, prolonged the action potential in about 75% of the neurons of dorsal root ganglia in ganglion spinal cord explants of mouse (tested in 5 mM Ba 2+). Both the excitatory and inhibitory effects of these kappa opioids were prevented by perfusion together with the opioid antagonist, diprenorphine (10nM). However, when responsivity tests with opioids were carried out in the presence of multiple K + channel Mockers [Ba 2+, Cs + and tetraethylammonium (TEA)], 1 nM dynorphin prolonged the action potential in only 7% of the neurons ( n = 28), whereas 1 nM U-50,488H still elicited the prolongation of the action potential in 60% of the cells ( n = 39). These data suggest that dynorphin prolongs the action potential of neurons of dorsal root ganglion by activating a kappa subtype of receptor that decreases a voltage-sensitive K + conductance, whereas U-50,488H produces similar excitatory modulation of the action potential by activating another kappa subtype of receptor that increases a voltage-sensitive Ca 2+ conductance. Thus, U-50,488H-induced prolongation of the action potential appears to be mediated by a kappa subtype of receptor that produces the opposite effect on Ca 2+ channels to that which occurs during kappa opioid-induced shortening of the action potential. On the other hand, dynorphin-induced prolongation of the action potential is mediated by a kappa opioid-receptor that decreases a voltage-sensitive K + conductance. After chronic exposure to d-ala 2- d-leu 5 enkephalin (DADLE), neurons of dorsal root ganglia became tolerant to dynorphin-induced excitatory, but not inhibitory, modulatory effects on the action potential. Electrophysiological studies of multiple modes of kappa opioid modulation of the action potential of dorsal root ganglion neurons in vitro may provide clues to the functional roles of high- and low-affinity kappa receptors, that have been reported in opioid binding assays of spinal cord and brain tissues, and may help to analyze the second messengers that couple subtypes of kappa receptors to K + and Ca 2+ channels.

O-090 - Changes in the brain Met-enkephalin, Leu-enkephalin and dynorphin concentrations in stressful situations

O-090 - Changes in the brain Met-enkephalin, Leu-enkephalin and dynorphin concentrations in stressful situations