Endogenous descending modulation: spatiotemporal effect of dynamic imbalance between descending facilitation and inhibition of nociception.

In conscious rats, we investigated the change of nociceptive paw withdrawal reflexes elicited by mechanical and heat stimuli during intramuscular (i.m.) 5.8% hypertonic (HT) saline elicited muscle nociception. i.m. injection of HT saline caused rapid onset, long lasting (around 7 days), bilateral mechanical hyperalgesia, while it induced bilateral, slower onset (1 day after the HT saline injection), long-term (about 1-2 weeks) heat hypoalgesia. Ipsilateral topical pre-treatment of the sciatic nerve with 1% capsaicin significantly prevented the occurrence of both the bilateral mechanical hyperalgesia and the contralateral heat hypoalgesia. Intrathecal administration of either 6-hydroxydopamine hydrobromide (6-OHDA) or 5,7-dihydroxytryptamine (5,7-DHT), and intraperitoneal injection of naloxone all markedly attenuated the HT saline induced bilateral heat hypoalgesia, but not the mechanical hyperalgesia. Combined with experiments with lesioning of the rostroventral medulla with kainic acid, the present data indicate that unilateral i.m. injection of HT saline elicits time-dependent bilateral long-term mechanical hyperalgesia and heat hypoalgesia, which were modulated by descending facilitatory and inhibitory controls, respectively. We hypothesize that supraspinal structures may function to discriminate between afferent noxious inputs mediated by Aδ- and C-fibres, either facilitating Aδ-fibre mediated responses or inhibiting C-fibre mediated activities. However, this discriminative function is physiologically silent or inactive, and can be triggered by stimulation of peripheral C-fibre afferents. Importantly, in contrast to the rapid onset of descending facilitation, the late occurrence of descending inhibition suggests a requirement of continuous C-fibre input and temporal summation. Thus, a reduction of C-fibre input using exogenous analgesic agents, i.e. opioids, may counteract the endogenous descending inhibition.

Serotonin analog selectively ablates indentified neurons in the leech embryo.

Exposure of embryonic leeches to 5,7-dihydroxytryptamine a cytotoxic analog of the monoamine neurotransmitter serotonin, results in the selective ablation of serotonin-containing neurons in the ventral nerve cord. Other neurons appear to be unaffected by this treatment, including those that contain another monoamine neurotransmitter, dopamine. Embryos with ablations continue to develop into juvenile leeches, but as juveniles they are unable to make normal swimming movements. However, normal swimming movements can be instated in such leeches by injecting them with serotonin.

Localization of the serotonergic terminal fields modulating seizures in the genetically epilepsy-prone rat.

Serotonin (5-HT) has been shown to exert antiepileptic effects in a variety of generalized convulsive seizure models, particularly the genetically epilepsy-prone rat (GEPR). The present study was designed to identify the region/site(s) where 5-HT exerts anticonvulsant effects in the GEPR-9, a model in which sound-evoked generalized tonic-clonic seizures (GTCS) are highly sensitive to manipulations in 5-HT concentration. Because the 5-HT reuptake inhibitor, fluoxetine, was known to exert anticonvulsant effects in GEPR-9s via a 5-HT-dependent mechanism, we utilized selective regional 5-HT depletion in combination with systemic fluoxetine administration to find the site where a 5-HT deficit would prevent the anticonvulsant action of fluoxetine. Widespread destruction of serotonergic terminal fields or regionally specific terminal field destruction was achieved using intracerebroventricular and more target specific infusions of 5,7-dihydroxytryptamine. The capacity of fluoxetine to suppress seizures in GEPR-9s following a loss of 5-HT was then examined. The present findings show the anticonvulsant action of fluoxetine is markedly attenuated following the loss of midbrain 5-HT, particularly in the region of the superior colliculus, while forebrain and spinal cord 5-HT do not appear to play a role in the action of fluoxetine. The importance of the deep layers of the SC was confirmed by demonstrating that direct microinfusion of fluoxetine into the SC can suppress seizures in rats pretreated with the 5-HT(1A) receptor antagonist pindolol.

Serotonergic modulation of septo-hippocampal and septo-mammillary theta activity during spatial learning, in the rat.

Theta activity has been related to the processing of spatial information and the formation of hippocampus-dependent memory. The medial septum (MS) plays an important role in the control and coordination of theta activity, as well as in the modulation of learning. It has been established that increased serotonergic activity may desynchronize theta activity, while reduced serotonergic activity produces continuous and persistent theta activity in the hippocampus. We investigate whether serotonin acting on the medial septum could modify spatial learning and the functional relationship between septo-hippocampal and septo-mammillary theta activity. The serotonin was depleted (5HT-D) from the medial septum by the injection of 5,7 DHT (5,7- dihydroxytryptamine). Theta activity was recorded in the dorsal hippocampus, MS and mammillary nuclei (SUM, MM) of Sprague-Dawley male rats during spatial learning in the Morris water maze. Spatial learning was facilitated, and the frequency of the hippocampal theta activity during the first days of training increased (to 8.5Hz) in the 5HT-D group, unlike the vehicle group. Additionally, the coherence between the MS-hippocampus and the MS-mammillary nuclei was higher during the second day of the test compared to the vehicle group. We demonstrated that septal serotonin depletion facilitates the acquisition of spatial information in association with a higher functional coupling of the medial septum with the hippocampus and mammillary nuclei. Serotonin, acting in the medial septum, modulates hippocampal theta activity and spatial learning.

Characterization of the inducible serotonin-sensitive dihydroalprenolol binding sites with low affinity for isoproterenol.

The previous findings that the inducible [3H]-dihydroalprenolol (DHA) binding sites with low affinity for isoproterenol (RL) could be regulated by serotonin (5-HT) in vitro and by 5-hydroxytryptophan and the 5-HT uptake inhibitor fluoxetine in vivo, prompted the present pharmacologic characterization of these receptor sites, using nonlinear regression analysis of competition binding curves. If isoproterenol was used as the displacing agent, lesioning with 5,7-dihydroxytryptamine selectively increased [3H]-DHA binding sites with low micromolar affinity. By contrast, if 5-HT was used as the displacing agent, the receptor population with high agonist affinity showed a fourfold increase whereas the density of [3H]-DHA sites with low micromolar affinity for 5-HT was not altered. Neither the 5-HT1A agonist, 8-OH-DPAT, nor mianserin, a 5-HT2 and 5-HT1C antagonist, altered the induced RL receptor population, whereas the selective 5-HT1B agonist CGS-12066B reduced the increase in the RL receptor population with a potency equal to that of 5-HT. These results strengthen the notion that the [3H]-DHA sites with low agonist affinity for isoproterenol represent 5-HT1B receptors induced following a reduction of serotonergic neuronal function.

Intracerebroventricular administration of 5,7-dihydroxytryptamine to mice increases both head-twitch response and the number of cortical 5-HT2 receptors.

5-Hydroxytryptamine-containing (5-HT) neurones in brain of the mouse were selectively destroyed by intracerebroventricular injection of 5,7-dihydroxytryptamine (5,7-DHT, 50 micrograms). Sham-lesioned controls received vehicle (2 microliters, i.c.v.). Two weeks later the head-twitch response induced by 5-methoxy-N,N-dimethyltryptamine (2.0 mg/kg) and mediated by 5-HT2 receptors was markedly enhanced in the lesioned mice. Furthermore, lesioning also increased 5-HT2 binding sites in the cortex, assessed by the binding of [3H]ketanserin in these animals, and decreased levels of 5-HT in the cortex (70%) and mid/hindbrain (64%) regions. A second group of mice, lesioned with less 5,7-DHT (5-20 micrograms, i.c.v.) showed unaltered head-twitch responses to 5-methoxy-N, N-dimethyltryptamine (2.0 mg/kg) and did not have increased 5-HT2 receptor binding in the cortex. Depletions of 5-HT were 32 and 40% in the cortex and mid/hindbrain, respectively. Comparison of the extent of depletion of 5-HT in the mid/hindbrain after lesioning, with the increase in head-twitch response, suggested that depletions of less than 40% did not affect this behaviour. However, depletions greater than 40% produced marked increases in this response and there was a good correlation between these two variables (r=0.701, P less than 0.01). In conclusion, these data suggest that enhanced head-twitch responses occurring after lesioning with 5,7-DHT may result from increased 5-HT2 receptors in brain. Moreover, the magnitude of the observed behavioural enhancement was dependent upon the extent of depletion of 5-HT produced by the lesioning.

The effects of 5,7-dihydroxytryptamine and p-chlorophenylalanine on thyrotrophin-releasing hormone in regions of the brain and spinal cord of the rat.

The distribution of thyrotrophin-releasing hormone (TRH) and 5-hydroxytryptamine (5-HT) were compared in ten regions of the rat brain and in lumbar spinal cord. After dissection, using a cutting box and tissue punches, TRH was measured by radioimmunoassay and 5-HT by HPLC with electrochemical detection. Within the brain the highest levels of TRH were found in the median eminence and the remaining hypothalamus. There were also relatively high levels in the suprachiasmatic nucleus, septal nuclei and nucleus accumbens. Highest levels of 5-HT were found in the raphe nuclei, hypothalamic nuclei, nucleus accumbens, stria terminalis, septal nuclei and hippocampus. 5,7-Dihydroxytryptamine (5,7-DHT; 200 micrograms, i.c.v.) markedly reduced levels of 5-HT in brain and spinal cord. In the ventral lumbar cord there was a comparable decrease of TRH and 5-HT (-80%) and a smaller but significant decrease in the nucleus accumbens (-55%) and septal nuclei (-38%). p-Chlorophenylalanine (PCPA; 250 mg/kg X 2) reduced levels of 5-HT (-80%), without significantly altering those of noradrenaline. p-Chlorophenylalanine also significantly reduced levels of TRH in the nucleus accumbens (-72%) but not in the other regions of brain or spinal cord taken. The results are discussed in relation to the previously described co-existence of TRH and 5-HT in the spinal cord and the possible alternative forms of interactions between amine and peptide in the nucleus accumbens.

The effects of monoamine neurotoxins on peptides in the rat spinal cord.

The coexistence of two neuronally-localised peptides, substance P and thyrotropin-releasing hormone (TRH), in descending serotoninergic nerve fibres to the spinal cord was investigated using immunocytochemical and biochemical methods. Substance P-like material in the spinal cord was shown to be identical to the undecapeptide substance P by the criteria of gel filtration, high performance liquid chromatography and behaviour in substance P specific radioimmunoassays. Immunocytochemical staining for 5-hydroxytryptamine, substance P, and TRH showed that all three substances had a similar distribution in nerve fibres and terminals in the ventral and lateral grey matter of the spinal cord. After treatment with the serotonin neurotoxin 5,7-dihydroxytryptamine, neuronal elements containing 5-hydroxytryptamine, substance P and TRH degenerated and disappeared from these parts of the spinal cord in parallel with one another. Biochemical measurements of 5-hydroxytryptamine, substance P and TRH in the spinal cord after treatment with 5,7-dihydroxytryptamine confirmed that these three substances were all depleted from the ventral horn and, in addition, showed that there was a small depletion of substance P from the dorsal horn. Two other neuropeptides, somatostatin and methionine-enkephalin were not depleted from the spinal cord by treatment with 5,7-dihydroxytryptamine nor was substance P in other parts of the brain. Substance P in the spinal cord was unaffected by 6-hydroxydopamine, a drug known to destroy catecholamine-containing neurones. These results are consistent with coexistence of substance P and TRH together with 5-hydroxytryptamine in the descending axons and terminals of bulbospinal neurones.

Effect of intracisternal 5,-7-dihydroxytryptamine on the acute antihypertensive action of propranolol in the sino-aortic denervated anaesthetized dog.

1 The anti-hypertensive effects of intravenously and intracisternally administered (+/-)-propranolol were studied in anaesthetized dogs with acute neurogenic (sino-aortic denervation) hypertension. The animals were pretreated 7 days earlier with intracisternally administered 5,7-dihydroxytryptamine (5,7-DHT 200 microgram/kg plus desipramine 5 mg/kg i.v.). 2 5,7-DHT (plus desipramine) failed to decrease both basic blood pressure and heart rate measured before sino-aortic denervation. After 5,7-DHT (plus desipramine) pretreatment, acute sino-aortic denervation induced a rise in blood pressure and stimulated the heart rate, these effects being similar (in intensity and duration) to those observed in control (saline-pre-treated) debuffered dogs during the first hour following the deafferentation. 3 In debuffered dogs, (+/-)-propranolol given by intracisternal (50 microgram/kg) or intravenous (300 microgram/kg) routes decreased both blood pressure and heart rate. 4 5,-DHT (plus desipramine) pretreatment abolished the antihypertensive effect of intracisternal propranolol whereas the action of intravenous propranolol was only delayed. In contrast, this pretreatment failed to reduce and even sometimes enhanced the negative chronotropic response induced by propranolol. 5 These results suggest that central 5-hydroxytryptaminergic pathways play an important role in the acute hypotension elicited by intracisternal (+/-)-propranolol in debuffered hypertensive anaesthetized dogs, but little, if any in propranolol-induced bradycardia.

Increased tryptophan hydroxylase activity in serotonergic nerve terminals spared by 5,7-dihydroxytryptamine.

Adult rats received intraventricular injections of 5,7-dihydroxytryptamine (5,7-DHT) to destroy serotonin (5-HT)-containing nerve terminals throughout the brain. When the animals were killed 3 or 21 days later, we observed a marked decrease in 5-HT content in septum and hippocampus and a parallel decline in in vitro high affinity 5-HT uptake. 5-Hydroxyindoleacetic acid (5-HIAA) concentrations also were reduced but by a much smaller extent, resulting in significant increases in the ratio of 5-HIAA to 5-HT. These changes were accompanied by similar increases in the ratio of tryptophan hydroxylase (TPH) activity to 5-HT content. The relative increases in TPH activity resulted from two temporally distinct processes, the first of which appeared to be an activation that could be mimicked in vitro by Ca2+-dependent phosphorylation. We conclude that, after partial damage to 5-HT neurons, there is a compensatory increase in the synthesis and release of 5-HT from those terminals that remain.

Pharmacological analysis of the neurotransmitter mechanisms regulating phenylethanolamine N-methyltransferase in the adrenal gland.

The i.p. administration of reserpine daily for 4 days to rats brought about an increase of adrenal phenylethanolamine N-methyltransferase (PNMT) activity. However, the combination of the systemic administration of p-chlorophenylalanine (PCPA) and reserpine for 3 days produced an earlier increase in this adrenal enzyme. The effect was reduced significantly in the denervated gland. Prior administration of 5,7-dihydroxytryptamine (DHT) i.c.v. to rats greatly potentiated the inducing effect of reserpine. On the other hand, the depletion of catecholamines by giving rats alpha-methyl-p-tyrosine (AMPT) i.p. or 6-hydroxydopamine (6-OHDA) i.c.v. did not alter the action of reserpine on adrenal PNMT. PCPA, DHT, AMPT and 6-OHDA did not have any effect by themselves on adrenal PNMT, but the combination of PCPA and AMPT, each given i.p., caused increased adrenal PNMT activity. The administration of dopamine agonists, a treatment that increases adrenal TH, did not modify adrenal PNMT. We conclude that the induction of PNMT by reserpine involves depletion of catecholamines and serotonin, the depletion of serotonin having the more powerful effect. A monoaminergic (serotonergic) inhibitory pathway is involved in the central regulation of adrenal PNMT activity.

Hippocampal cell nuclear binding of corticosterone following 5,7-dihydroxytryptamine.

Adult male rats were injected into the lateral brain ventricle with 5,7-dihydroxytryptamine (5,7-DHT). They were adrenalectomized 5-7 days later and, following an additional 24 h, the specific in vitro [3H]corticosterone binding capacity of dorsal hippocampal slices was determined by estimation of uptake of radioactivity by the nuclear fraction. Specific corticosterone (CS) binding was reduced by 50-70% in the neurotoxin-treated as compared to vehicle-injected animals. Brain serotonin and 5-hydroxyindoleacetic acid concentrations were depleted by 50-70% in the 5,7-DHT-injected rats. These results suggest that the maintenance of normal dorsal hippocampal CS binding capacity is dependent upon the integrity of endogenous brain serotoninergic neuronal systems.

Effects of neonatal administration of 5,7-dihydroxytryptamine on locomotor activity.

Neonatal rats treated on day 3 of life with 50 or 100 micrograms 5,7-dihydroxytryptamine exhibited long-lasting selective depletions of serotonin (5-HT). The 5-HT depletions produced a shift in the peak in locomotor activity from its normal occurrence at 15 days of age to later days of age. The observation that the decreases in activity after the peak were delayed, rather than eliminated, suggests that the inhibition of locomotor activity produced by 5-HT may be of transient importance in the developing rat. The transience of the inhibition may be the result of the continuing development of nonserotonergic systems during this time period that are involved in the regulation of activity.

Effects of 5,7-dihydroxytryptamine and 6-hydroxydopamine on head-twitch response induced by serotonin, p-chloroamphetamine, and tryptamine in mice.

Head-twitch response (HTR) in mice was induced by intracerebroventricular injection of tryptamine (TRA) as well as serotonin (5-HT) and p-chloroamphetamine (PCA). Pretreatment with 5,7-dihydroxytryptamine enhanced both the 5-HT-induced and the TRA-induced HTR. The PCA-induced HTR, however, was attenuated by the drug. On the other hand, pretreatment with 6-hydroxydopamine did not alter the 5-HT response but enhanced both the PCA- and the TRA-induced response. These results suggest that 5-HT may directly stimulate the post-synaptic receptors, while the PCA response may be based on the release of endogenous 5-HT. The presynaptic component of the central serotonergic system does not appear to be involved in the TRA response. Both PCA and TRA may affect catecholaminergic systems which can suppress the response.

Evidence against the involvement of serotonergic neurons in the anti-punishment activity of diazepam in the rat.

The effects of manipulating central serotonergic transmission were assessed on the anti-punishment effects of diazepam (2 mg/kg IP) in rats. In a paradigm involving the inhibition of pressing for food induced by the delivery of a signal previously associated with electric foot-shocks, lesioning serotonergic neurons of the dorsal raphé with the neurotoxin 5,7-dihydroxytryptamine (5,7-DHT; 1 microgram in 0.4 microliter) neither affected behavioral inhibition in control rats nor modified the ability of diazepam to release responding. Furthermore, suppression of pressing for food induced by a fixed ratio 7 schedule of shock presentation was reduced by bilateral infusion of 5,7-DHT (2 micrograms in 0.5 microliter) into the substantia nigra, but the ability of diazepam to increase punished responding was preserved. Finally, blockade of benzodiazepine-induced decrease in serotonin release by application of the benzodiazepine receptor antagonist Ro 15-1788 (10(-5)-10(-4)M in 0.2 microliter) into the dorsal raphé did not alter the releasing effect of diazepam on suppression of pressing for food caused by a signal of punishment. At these concentrations. Ro 15-1788 was devoid of any effect on behavioral inhibition in control rats. Taken together, these results indicate that the anti-punishment activity of benzodiazepines can be dissociated from the reduction in tryptaminergic transmission produced by these drugs.

Facilitation of shock-induced fighting following intraventricular 5,7-dihydroxytryptamine and 6-hydroxydopa.

Using a 15-s intershock interval, an increase in shock-induced fighting was observed following intraventricular 96 microgram 5,7-dihydroxytryptamine (5,7-DHT) and 90 microgram 6-hydroxydopa (6-OHdopa). The incidence of predatory mouse killing was enhanced by 5,7-DHT, but was not affected by 6-OHdopa. Pain sensitivity was increased by 6-OHdopa, but both neurotoxins produced hyperreactivity to footshock. Specific serotonin depletion was produced by 5,7-DHT and norepinephrine depletion by 6-OHdopa. The increase in shock-induced fighting could not be predicted on the basis of monoamine depletion alone, since a long intershock interval was necessary to observe this increase.

Enhanced selective 5-HT depletions in the DHT rat model: denervation supersensitivity and recovery of function.

The effects of enhancing 5-HT depletion with multiple intracisternal injections of 5,7-dihydroxytryptamine (DHT) on spontaneous or L-5-hydroxytryptophan (5-HTP)-induced behaviors (videotaped) and locomotor activity (photocell recording) were studied in the adult rat. After four DHT injections, 5-HT content in septum/accumbens, hippocampus, striatum, neocortex, cerebellum, and cervical spinal cord fell to 0-10% of controls. Multiple injections also significantly improved depletions in brainstem and diencephalon, which were not as extensive. Spontaneous locomotor activity (LMA) was increased in DHT-lesioned rats for 1 week. The associated behavioral abnormalities, hindlimb hyperextension and incomplete rearing were also transient and differed from the motor syndrome evoked by 5-HTP. Multiple DHT injections did not qualitatively modify the 5-HTP syndrome but shifted the dose response curve to the left compared to single injections. Syndrome behaviors shared a similar dose threshold and could be evoked with 30 mg/kg 5-HTP. Two weeks after DHT, the locomotor response to 5-HTP (65 mg/kg) was method dependent or biphasic: decreased in brief recordings when syndrome abnormalities were greatest and increased in hour-long recordings. LMA correlated with rearing in controls and inversely with total behavioral abnormality in DHT-lesioned rats injected with 5-HTP. Multiple regression of LMA with regional 5-HT content was significant for hippocampus, striatum, and septum/accumbens. These data suggest that the development of denervation supersensitivity, the proposed mechanism of the 5-HTP-evoked motor syndrome, may be responsible for the rapid recovery of function in LMA.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of depletion of brain 5-hydroxytryptamine by 5,7-dihydroxytryptamine on ethanol tolerance and dependence in the rat.

Brain 5-hydroxytryptamine (5-HT) was depleted in rats by intraventricular injection with 5,7-dihyroxytryptamine (5,7-DHT) prior to feeding rats a liquid diet containing ethanol. After withdrawal of ethanol, withdrawal reactions were significantly less severe in 5-HT-depleted rats than control rats. Sleeping times after a standard dose of ethanol or pentobarbitone were significantly prolonged in 5-HT-depleted rats. However, metabolic and pharmacodynamic tolerance developed to a similar extent in 5-HT-depleted rats as in control rats. It was concluded that 5-hydroxytryptaminergic neurons are not directly involved in the development of physical dependence on or tolerance to ethanol. Depletion of brain 5-HT by 5,7-DHT appears to result in a non-specific central nervous system depression that potentiates the depressant actions of ethanol and pentobarbitone and antagonises the hyperexcitability of ethanol withdrawal.

Effect of 5,7-dihydroxytryptamine on the development of tolerance to ethanol.

5,7-Dihydroxytryptamine (5,7-DHT) or the vehicle was administered once into both lateral ventricles of the rat. Desmethylimipramine (DMI) was administered IP prior to the intraventricular injection of 5,7-DHT to prevent the destruction of norepinephrine (NE) terminals. Following recovery from surgery, ethanol (5 g/kg, PO) or isocaloric sucrose was given daily for 25 days. Tests at 5-day intervals showed that chronic ethanol treatment produced tolerance to the motor impairment on the moving belt test and to hypothermic effects of ethanol. The 5,7-DHT treatment did not alter either the motor impairment or hypothermia produced by the initial dose of ethanol. However, 5,7-DHT treatment produced a 75% depletion of brain serotonin (5-HT) without altering NE concentration and retarded the development of tolerance to ethanol in both measurements. This study with a specific central depletor of 5-HT, without alteration in NE concentration, extends and supports our hypothesis that brain 5-HT modulates the development of tolerance to ethanol.

Supersensitivity to L-5-hydroxytryptophan after 5,7-dihydroxytryptamine injections in desmethylimipramine- and nomifensine-pretreated rats: behavioral evidence for postsynaptic supersensitivity.

The behavioral syndrome induced by L-5-hydroxytryptophan (L-5-HTP) in rats was used to study the supersensitivity to L-5-HTP and 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) which develops after unilateral intracerebroventricular (ICV) injections of 200 microgram 5,7-dihydroxytryptamine (5,7-DHT). Pretreatment of the animals with a combination of desipramine and nomifensine was found to protect dopamine neurones better than desipramine alone. Maximal behavioral supersensitivity to L-5-HTP and 5-MeODMT was found as early as 24 h after injection of the neurotoxin, even in the presence of the specific 5-HT uptake inhibitor CGP 6085 A, or the MAO-A inhibitor clorgyline. The results indicate that a quickly occurring postsynaptic event contributes to the development of behavioral supersensitivity after ICV injections of 5,7-DHT.