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Alteration of metabolic activity during deep halothane anesthesia (3%) was analyzed qualitatively in the rat using a modified 2-deoxyglucose technique using the tritiated compound. The grey matter exhibited a homogeneous low level of metabolic activity in most places but some nuclei or subnuclear structures showed a much higher activity. These included in particular the locus coeruleus, the dorsal raphé, the substantia nigra pars compacta, the CA3 region of the hippocampus and the nucleus reticularis thalami. In the nucleus reticularis thalami, the use of 2-deoxy-[3H]glucose allowed us to demonstrate a high cellular metabolic activity, which is in agreement with several electrophysiological studies which showed a high rate of spontaneous activity under the same anesthetics.

1. It has previously been shown that, in normal humans, heterotopic painful thermal conditioning stimuli induce parallel increase in the thresholds of a spinal nociceptive flexion reflex (RIII reflex) and the concurrent sensation of pain elicited by electrical stimulation of the sural nerve. On the basis of analogous animal studies, we proposed that such phenomena could be related to diffuse noxious inhibitory controls (DNIC), which have been described in the rat. The present study, which was carried out on normal volunteer subjects, was particularly concerned with the extent and temporal characteristics of the depressive effects of DNIC triggered by painful thermal stimuli on RIII reflex activity. In addition, because it was possible that these depressive effects could have resulted from a direct postsynaptic inhibition of motoneurons, a second part of the study was aimed at determining whether or not the heterotopic noxious thermal stimuli also affected the excitability of alpha-motoneurons, as monitored by the monosynaptic Hoffmann reflex (H reflex) technique. 2. In the 11 subjects under study, application of moderate, nonnoxious temperatures (40-44 degree C) to the contralateral hand (via a thermoregulated and agitated waterbath) did not modify the RIII reflex nor the associated sensation of pain. By contrast noxious temperatures clearly depressed the RIII reflex and the concurrent sensation of pain, both during and after the conditioning procedure (CP), in a direct linear relationship to the temperature of the waterbath in the 45-47 degree C range; the maximal depressive effect was observed with the highest conditioning temperature. A significant relationship was also found between the extent of the RIII depression during the CP and that during a 10-min period of post-CP observation. 3. The depressive effects observed on both the RIII reflex and pain were not associated with clear change in autonomic functions. Respiration remained stable during the sessions, with no significant relationship between the temperatures of the waterbath and respiratory rate. Heart rate was slightly but significantly increased during the immersion of the hand in the 46 or 47 degree C waterbaths; this increase, however, ceased with the end of CP. 4. Application of thermal conditioning stimuli produced a slight but nonsignificant increase of the monosynaptic H reflex during the first minute of CP, no matter what was the temperature of the waterbath. However, there were no subsequent variations during the 6-min period of post-CP observation.(ABSTRACT TRUNCATED AT 400 WORDS)

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.

The nociceptive flexion reflex (RIII reflex) and the concurrent subjective pain score elicited by right sural nerve stimulation at random intensities were studied in 10 healthy volunteers. A close relationship was found between the recruitment curves of the reflex and the pain score as a function of stimulus intensity. As a consequence, the threshold of the RIII reflex (Tr) and of pain sensation (Tp) were found to be almost identical (mean: 9.8 and 11.3 mA, respectively). Similarly, the threshold for obtaining a maximal reflex response (Tmr) was found to be very close to that for intolerable pain (Tip): 33.5 and 35.1 mA, respectively. These four parameters were studied before and during the immersion of the left hand into a heated thermoregulated waterbath at various temperatures (from 40 to 47.5 degrees C). While nonnociceptive temperatures (40 to 44 degrees C) were without effect, higher conditioning temperatures induced an increase in the four thresholds. In addition, a highly significant linear relationship was observed between the increase in these thresholds and the intensity of the conditioning stimulus in the 44 to 47.5 degrees C range. These four parameters were also studied before and during three other nociceptive conditioning stimuli: immersion of the left hand into a 6 degrees C waterbath, 10 watts muscular exercise of the left hand performed under ischaemia and a painful (5.5 kg/cm2) pinch applied on the nasal septum. These three conditioning situations induced a very significant increase of the four thresholds considered in this study with the greatest being observed during nociceptive cold applied to the left hand. During all the conditioning situations, variations in Tr and Tp as well as in Tmr and Tip were found to be linearly related. This indicates a close relationship between the effects of the conditioning nociceptive stimuli on the reflex and the related pain sensation. These results suggest that the modulation of pain by heterotopic nociceptive stimuli can be explained at least in part by a depression in the transmission of nociceptive messages at the spinal level. They are discussed with reference to the counterirritation phenomena and common features with 'diffuse noxious inhibitory controls' (DNIC) are underlined.

1. In order to study descending influences of the brain stem upon the transmission of nociceptive messages at the spinal level, the activities of lumbar lamina V dorsal horn cells, induced by intra-arterial injection of brandykinin into the limbs, were recorded in unanaesthetized cats in both decerebrate and temporary spinal states (reversible cold block applied at the thoracic level). 2. In the decerebrate state, the intra-arterial injection of bradykinin had little or no effect. 3. During the reversible spinalization, the effects of bradykinin were revealed or considerably enhanced. As described in a previous study, in the C1-transected cat, three types of effects were encountered: excitatory, inhibiitory and mixed (inhibitory-excitatory). 4. These modifications observed after spinalization were generally associated with a large increase of the spontaneous firing rate. 5. These results emphasize, in the decerebrate cat, the importance of descending inhibitory controls exerted by the brain stem upon the transmission of nonciceptive messages at the spinal cord level.

The relationship between stimulus intensity and its efficacy in inducing diffuse noxious inhibitory controls (DNIC) was investigated in anaesthetized rats by using thermal stimulation of the tail for conditioning dorsal horn convergent neuronal responses to C fibres emanating from the hindpaw extremity. The threshold for obtaining inhibition of the neuronal responses ranged between 40 and 44 degrees C and there was a highly significant correlation between noxious temperatures (44-52 degrees C) and the degree of inhibition. These data provide support for the notion of an involvement of inhibitory processes (DNIC) in the signalling of pain by convergent neurones.

Information concerning cell-wall associated proteinases of lactobacilli is limited. In Lactobacillus casei and Lactobacillus plantarum , presence of such proteinase is clearly shown. Differences between several strains were noticed. Higher cell-wall-associated proteinase activity can be measured in extracts obtained from milk-grown cells when compared to MRS-grown cells. No aminopeptidase, dipeptidase or carboxypeptidase activities were detected in the cell-wall-associated proteinase fraction. Isoelectric focusing of α s1 -casein hydrolysates obtained by the action of this fraction from L. casei grown in milk revealed the presence of a major hydrolysis product and three minor degradation products with isoelectric points more acidic than α S1 . Beta-casein was also degraded by the cell-wall extract with formation of one major product and several minor products with isoelectric points more acidic than β-casein. Two major hydrolysis products with isoelectric points higher than β-casein were also detected. Isoelectric focusing of α s1- and β-casein hydrolysates obtained by the action of the intracellular extracts of L. casei grown either in milk or in MRS broth shown identical patterns. As with L. casei , two strains of L. plantarum exhibited cell-wall proteinase activity. Milk-grown cells were more proteolytic than MRS-grown cells. Generally L. plantarum was significantly less proteolytic than L. casei .

The tripeptide Tyr-Gly-Gly (YGG), representing the product of enkephalin hydrolysis by enkephalinase (EC 3.4.24.11), was characterized and its levels measured in spinal cord perfusates of halothane-anaesthetized rats. During noxious pinching of the muzzle, which is known to trigger enkephalin release, YGG levels were enhanced more markedly and for longer than were those of [Met5]enkephalin (YGGFM), in the same samples. By contrast, neither YGG nor YGGFM levels were affected by pinching the tail. Treatment with carbaphethiol, a parenterally-active aminopeptidase inhibitor, markedly increased YGG levels and lengthened the duration of the increase produced by pinching the muzzle. Treatment with acetorphan, a parenterally-active enkephalinase inhibitor, given alone or in combination with carbaphethiol, completely prevented the rise in YGG triggered by noxious stimulation. By contrast, [Met5]enkephalin levels in the perfusates were increased by the combined administration of the two peptidase inhibitors but these levels were not further enhanced by noxious stimulation. Thus, spinal cord YGG appears to be formed under the influence of enkephalinase and to constitute a sensitive index of enkephalin release.

Nineteen convergent neurones, 19 'non-noxious only' neurones and 13 'proprioceptive' neurones all with peripheral excitatory receptive fields on the ipsilateral hindpaw, were recorded in the lumbar dorsal horn of non-spinalized, anaesthetized rats. These neurones were all excited by the electrophoretic application of glutamate; using 20 s applications of appropriate electrophoretic currents, almost identical levels of activity (around 30 spikes/s) were produced for each of the 3 types of neurones. The application of heterotopic noxious stimuli resulted in strong inhibitions of the glutamate-evoked activity of the convergent neurones. During the application of noxious heat (52 degrees C) to the tail and noxious pinches to the tail, contralateral hindpaw and muzzle, the glutamate-evoked activity was depressed by 78%, 84%, 63% and 59%, respectively. It was also found that these inhibitions outlasted the period of conditioning stimulation by several minutes (post-effects). Twenty seconds after i.p. injection of bradykinin (20 micrograms) the glutamate-evoked activity was depressed by 56% and this effect also lasted for several minutes. The application of identical heterotopic noxious stimuli did not affect the glutamate-evoked activity of the 'non-noxious only' neurones or the 'proprioceptive' neurones. The application to convergent neurones, of doses of glutamate, which were very much larger than the threshold for firing, produced an intense discharge followed by a progressive decrease in spike amplitude and finally a blocking of the spike discharge. During such sequences, which are typical of excessive depolarization, the application of noxious conditioning stimuli (e.g pinch of the contralateral hindpaw or muzzle) resulted in recovery of the spike discharge. In several cases, this recovery long outlasted the period of conditioning noxious stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

The nature of stimuli that activate neurones located within the Subnucleus Reticularis Dorsalis (SRD) was determined. These neurones do not respond to visual, auditory or proprioceptive stimuli but are preferentially or even exclusively activated by noxious stimuli applied to any part of the body; only neurones of a sub-group have a predominantly contralateral activation. All the neurones were exclusively activated by stimulation of A delta or A delta and C afferent fibres. These observations suggest that this well-delimited reticular structure is specifically involved in the processing of nociceptive information.

Although the physiological functions of the endogenous opioid systems are not yet clearly established, it is widely accepted that they exert an inhibitory control on pain transmission. However, the well-documented hypoalgesic effects of low doses of the opiate antagonist naloxone both in animals and humans do not fit in with this concept. The present investigations, at two different spinal/medullary levels (viz. cervicotrigeminal and lumbar) demonstrate that, in the rat, a noxious mechanical stimulus does not alter the release of Met-enkephalin-like material (MELM) from neural segments related to the stimulated area of the body, but does increase its release from other segments. Electrophysiological studies have already demonstrated the existence of such heterosegmental mechanisms, notably 'diffuse noxious inhibitory controls' (DNIC), which are naloxone-reversible and could play an important role in pain perception. The involvement of spinal enkephalins in DNIC would seem to mean that the heterosegmental spinal release of MELM triggered by noxious stimuli participates in pain processes.

The analgesic effect of the microinjection of low doses of methione-enkephalin (20 micrograms in 0.5 microliter) into the caudal brain stem of the unrestrained rat was investigated by means of one vocalisation test. Immediate short duration significant increases in threshold (21%) were seen from sites in the nucleus raphé magnus (NRM). Delayed effects were seen from sites immediately adjacent to this nucleus; sites more lateral produced no significant effect. These results lend further support to the postulated role of NRM in antinociception.

Potent analgesia results from electrical stimulation of the periaqueductal grey matter in several species including man. Electrophysiological experiments indicate that this electrical analgesia could result from the activation of descending influences which inhibit the activity of dorsal horn interneurons in the transmission of painful messages. Numerous physiological, behavioral and pharmacological studies mention that the descending serotoninergic Bulbo-spinal system plays a major role in electrical analgesia. Several studies suggest that both electrical and morphine analgesia share, at least in part, a common site and mechanism of action. This possibility is mainly supported by the fact that analgesia induced by electrical stimulation is suppressed or reduced by a specific opiate antagonist (Naloxone).

15-(p-Iodophenyl) pentadecanoic acid labelled with carbon-14 in the carboxyl group is obtained with good yields in five steps from [14C]NaCN, starting from 14-bromo 1-phenyl tetradecane 2. Specific radioactivity: 0.019 mCi/mg, 8.45 mCi/mmol; radiochemical yield: 74.6%.

Recordings were made from convergent neurons in the lumbar dorsal horn of the rat. These neurons were activated by both innocuous and noxious stimuli applied to their excitatory receptive fields located on the extremity of the ipsilateral hindpaw. Transcutaneous application of suprathreshold 2-msec square-wave pulses to the center of the receptive field resulted in responses to A- and C-fiber activation being observed: 27.2 +/- 2.2 (mean +/- S.E.M.) C-fiber latency spikes were evoked per stimulus. This type of response was inhibited by applying noxious conditioning stimuli to heterotopic areas of the body; in particular, immersing the tail in a 52 degrees C waterbath caused a 74.2 +/- 2.0% inhibition of the C-fiber evoked responses; such inhibitory processes have been termed diffuse noxious inhibitory controls (DNIC). The effects of microinjections of morphine (0.6-40 micrograms; 2 microliter) within the 3rd ventricle on both the unconditioned C-fiber-evoked responses and the inhibitory processes triggered from the tail were investigated in an attempt to answer two questions: 1) does i.c.v. morphine increase tonic descending inhibitory processes? and 2) what are the effects of i.c.v. morphine on descending inhibitory processes triggered phasically by noxious stimuli? The predominant effect of i.c.v. morphine on the C-fiber-evoked responses was a facilitation (17 of 26 cases). Such a facilitation was dose-related in the 0.6 to 40 microgram range and naloxone reversible; it plateaued from 20 min after the microinjection. No clear relationship was found between the number of C-fiber evoked responses in the control sequences and the subsequent effect of i.c.v. morphine. Intracerebroventricular morphine clearly reduced DNIC in the majority of cases (21 of 26). Such a reduction was dose-related in the 0.6 to 2.5 microgram range and naloxone reversible; it plateaued within 90 min of microinjection. No clear relationship was found between the changes in DNIC and either the number of C-fiber-evoked spikes in the control sequences or the changes in the C-fiber responses induced by i.c.v. morphine. Autoradiographic controls using [3H]morphine showed a labeling along the ventricle wall including the hypothalamus, the periaqueductal gray matter and the floor of the 4th ventricle, three regions which have been implicated in the control of nociceptive transmission at the spinal level. Diffusion from the ventricle wall was over a distance of 0.5 mm and was identical whether observed 20 or 95 min after the microinjections.(ABSTRACT TRUNCATED AT 400 WORDS)

Recordings were made from dorsal horn neurones in the spinal cord and trigeminal nucleus caudalis of intact anaesthetized rats. These rats had been rendered polyarthritic by s.c. injection of Mycobacterium butyricum suspended in oil into the base of the tail. The experiments were carried out during the acute phase of the illness (3-4 weeks post inoculation) during which hyperalgesia occurred. The disease mainly affected the hind paws and the tail and, to a lesser extent, the forepaws. The facial area of the animals was not at all affected. As described in a previous paper, recordings from lumbar dorsal horn neurones revealed that two subpopulations could be described on the basis of their electrophysiological characteristics. Namely, 'typical' units which include convergent, non-noxious and proprioceptive neurones and which have properties essentially similar to those found in healthy rats, and 'atypical' cells which have no counterpart in healthy rats and which include convergent and non-noxious neurones. All the typical convergent neurones were inhibited by noxious stimuli applied to heterotopic body areas, whereas typical non-noxious and proprioceptive neurones were not; these observations are similar to those described in healthy rats as diffuse noxious inhibitory controls (DNIC). However, it was also found that 88% of the atypical convergent and 85% of the atypical non-noxious cells were inhibited by various heterotopic stimuli. The most important observation was that gentle stimulation such as mild pressure applied to the inflamed contralateral ankle joint--a stimulus intensity which has never been found to be effective in healthy animals--was capable of triggering inhibition of both typical and atypical convergent neurones. Recordings from trigeminal nucleus caudalis neurones revealed that the entire population presented essentially the same properties as those observed in healthy animals in terms of activity evoked by natural or electrical stimulation of their excitatory receptive fields. The activity of non-noxious neurones was never modified by any heterotopically applied stimuli. By contrast, all convergent neurones were inhibited by heterotopic stimuli, noxious (52 degrees C, pinch) or non-noxious (light and mild pressure), applied to inflamed areas. While the inhibition triggered by noxious stimuli was reminiscent of that observed in healthy rats, the inhibition triggered by non-noxious mechanical stimuli was related to the inflammatory state of the part of the body stimulated, the most sensitive areas being the hind paws.(ABSTRACT TRUNCATED AT 400 WORDS)

In the anaesthetized Rat, the entire population of dorsal horn convergent neurones is differentially affected by a noxious stimulus: while exciting the segmental pool, it strongly inhibits the remaining population. The inhibitory effects, which involve supraspinal mechanisms, are reduced to a great extent in parachlorophenylalanine pretreated animals. The role of raphé-spinal serotonergic pathways in nociception is discussed.

The course of adjuvant-induced arthritis was followed, over an 11-week postinoculation period, in Sprague-Dawley rats, using both clinical and behavioural methods of study. Clinical observations included body weight, diameters of radiocarpal and tibiotarsal joints and radiological analysis of forepaws, hindpaws and vertebrae. Behavioural observations included those of spontaneous behaviour-mobility, exploring, rearing, penis licking and scratching-and of pain-related tests-the foot-bend procedure and Randall-Selitto test. In general, the time course of the disease could be divided into 4 stages, viz. a preclinical, an acute, a postacute and a recovery stage. The preclinical stage (first week) was characterized by discrete radiological lesions of the forepaws and a slight increase in the threshold for struggle triggered by foot pressure, but, above all, by no change in the other parameters. The acute stage (weeks 2-4) was clearly defined by a pause in body weight gain and by dramatic behavioural changes: a lack of mobility and exploring behaviour, a profound increase in scratching behaviour and signs of hyperalgesia. During this period, hindpaw and forepaw joint diameters increased dramatically. The behavioural modifications peaked at the same time (week 4) as there was an acceleration in the radiological abnormalities (soft tissue swelling, demineralization and erosion of bone extremities and joint space narrowing) which had begun during week 3. During the postacute stage (weeks 5-8), body weight and behavioural reactions began to return towards control values, but joint diameters and radiological scores either continued to increase or remained constant. During the recovery stage (weeks 9-11), behavioural scores and body weight returned to control values while the mean joint diameters remained constant and there was a slight decrease in the radiological scores. In spite of the complexity of the model used in these studies, it appeared quite clear that the acute phase of the disease was characterized by the onset of clinical symptoms and dramatic changes in behaviour, all of which could be related to the occurrence of pain; signs of pain were still present, albeit weaker, in the postacute phase. Such observations, taken together, help to validate adjuvant arthritis as an experimental model of chronic pain in Sprague-Dawley rats.

Writhing behaviour induced by a viscero-peritoneal nociceptive stimulus (i.p. acetic acid) has been investigated during the development of arthritis experimentally induced in rats by s. c. injection of Mycobacterium butyricum; experiments were carried out at various (1-9 weeks) post-inoculation periods. In the most severe phase (2-5 weeks post-inoculation), when arthritis-induced hyperaesthesia was most pronounced, writhing behaviour was strongly reduced (80%). In the following period, the progressive decrease of hyperaesthesia was contemporaneous with a recovery of the writhing behaviour. We conclude that, in this chronic pain model, the behavioural reaction to a viscero-peritoneal nociceptive stimulus is impaired. Such heterotopic inhibitory processes could be relevant to some paradoxical clinical observations such as the masking of a pain by the experience of pain at another locus (i. e. counter-irritation phenomenon).

The involvement of serotonergic mechanisms in diffuse noxious inhibitory controls (DNIC) acting on dorsal horn convergent neurones has been studied in the anaesthetized rat. 35 neurones activated by transcutaneous electrical stimulation of their hindpaw receptive fields giving clear large A-fibre and C-fibre responses were recorded. These activities were conditioned by DNIC, evoked by either noxious heat applied to the tail or noxious pinch of the nose. Cinanserin (4 mg/kg i.v.) and metergoline (5 mg/kg i.v.), serotonin (5-HT) receptor blockers, strongly reduced the inhibitory effects of DNIC whilst having no significant effect on the non-conditioned responses. 5-Hydroxytryptophan, a precursor of 5-HT synthesis, significantly potentiated the effect of DNIC. These results indicate an important role for descending serotonergic pathways in DNIC. The functional role of this system is discussed.

1. In anesthetized rats recordings were made from neurons in the medulla caudal to the obex. In the medullary dorsal horn, typical trigeminal noxious specific, nonnoxious specific, and convergent neurons were found. In nucleus cuneatus, typical dorsal column units were recorded. In subnucleus reticularis dorsalis (SRD), recordings were made from neurons which exhibited convergence of nociceptive inputs from the entire body. In subnucleus reticularis ventralis (SRV) we recorded both from neurons with spontaneous activity that were either unaffected or inhibited by noxious stimuli applied to various parts of the body and from respiratory neurons. The present paper deals particularly with the nature of the stimuli that activated reticular neurons exhibiting nociceptive convergence. 2. Neurons with nociceptive convergence could be activated by mechanical, thermal, or chemical noxious stimuli applied to widespread areas of the body. By using percutaneous electrical stimulation, we found that they responded to the activation of peripheral fibers in the A delta- and C-range. Two neuronal subpopulations were defined according to the way in which SRD neurons responded to the electrical stimuli, namely: "neurons with total nociceptive convergence" (TNC) and "neurons with partial nociceptive convergence" (PNC). 3. The great majority (84%) of TNC neurons did not exhibit spontaneous activity and none of these neurons gave responses to heterosensory (flashes, whistle sounds) or proprioceptive stimuli. Most (88%) did not respond to any kind of innocuous cutaneous stimuli. By contrast, the entire population of TNC neurons responded to noxious mechanical, thermal, and visceroperitoneal stimuli. In the majority of cases (71%) long-lasting afterdischarges were observed following cessation of the application of the nociceptive stimulus. 4. All the TNC neurons responded to suprathreshold percutaneous electrical stimulation (2-ms duration) with two peaks of activation no matter which part of the body was stimulated. By stimulating two regions of the tail, 100 mm apart, we determined that the early and late peaks of activation were triggered by activities in peripheral fibers with mean conduction velocities of 10.8 +/- 0.5 and 0.74 +/- 0.05 (SE) m/s, respectively, i.e., A delta- and C-fibers. The mean thresholds for obtaining A delta-fiber components were found in the 0.4-0.7-mA range; the mean thresholds for obtaining C-fiber components were found in the 6-7.5- and 3-4-mA range for the face and the other parts of the body, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

The interaction between morphine analgesia and the heterotopic hypoalgesic effects induced by visceral pain was investigated using a behavioral approach in the rat. Both systemic morphine and a dull pain of peritoneal-visceral origin (induced by i.p. phenylbenzoquinone, PBQ) increase the threshold for vocalization to tail-shock. The question of the interaction between these two factors was assessed by administering morphine during the period of increase in threshold induced by PBQ. Not only did we find that morphine did not increase the threshold further, but, indeed, very significantly decreased it, with the more obvious effect being observed for a moderate dose (0.33 mg/kg, i.v.). It is concluded that morphine antagonizes the heterotopic hypoalgesic effects of visceral pain. These results do not support the hypothesis that morphine increases inhibitory controls of pain triggered by noxious inputs.

The effects of the novel mixed peptidase inhibitor, kelatorphan [N-(R)-3-(N-hydroxyaminocarbonyl-2-benzyl-1-oxopropyl)-L-alanine], were compared to those of a combination of the potent "enkephalinase" inhibitor thiorphan and the nonselective aminopeptidase inhibitor bestatin, on the catabolism of [3H]Met-enkephalin and on the release of endogenous Met-enkephalin by the rat spinal cord in vitro and in vivo. At 20 microM, kelatorphan almost prevented completely the degradation of exogenous [3H] Met-enkephalin by slices of the dorsal zone of the lumbar enlargement. Similarly, the addition of 20 microM kelatorphan to a [3H] Met-enkephalin-containing artificial cerebrospinal fluid superfusing the whole spinal cord of halothane-anesthetized rats efficiently protected the exogenous peptide from enzymatic degradation. In contrast, in the same in vitro and in vivo models, thiorphan (1 microM) or bestatin (20 microM) alone was inactive, and only their combination induced a significant protection of the exogenous peptide. In vitro and in vivo, kelatorphan (20 microM) increased markedly the spontaneous outflow of endogenous Met-enkephalin-like material as well as the peptide overflow due to K+-induced depolarization (in vitro and in vivo) or noxious stimulation (in vivo). Under similar conditions, thiorphan (1 microM) plus bestatin (20 microM) also enhanced the efflux of Met-enkephalin-like material, but generally to a lower extent than kelatorphan. Compared to thiorphan plus bestatin, kelatorphan exerts additional inhibitory effects on dipeptidylaminopeptidase activity and the present results could indicate that this enzyme also may be involved in the inactivation of extracellular Met-enkephalin at the spinal level in rats.

Diffuse Noxious Inhibitory Controls (DNIC) were investigated in anaesthetized intact rats, with or without p-chlorophenylalanine (pCPA) pretreatment. Dorsal horn convergent neurones responding to both noxious and non-noxious stimuli applied to their excitatory receptive field located on the distal part of the hindlimb, were recorded in the lumbar spinal cord. These cells received A alpha and C fibre inputs as shown by electrical stimulation of their receptive field. In control animals, the evoked responses to C fibre inputs could be strongly inhibited by various noxious stimuli applied to widespread areas of the body: the inhibitory effects induced by intraperitoneal administration of bradykinin, pinch applied to the tail or muzzle and noxious heat applied to the tail were of 77%, 87%, 83% and 61% respectively. Long-lasting post-effects were seen in most cases after cessation of the application of the conditioning stimulus. Pretreatment with pCPA (300 mg/kg, i.p., 3 days) resulted in a strong reduction of DNIC. The inhibitory effects induced by intraperitoneal administration of bradykinin, pinch applied to the tail or muzzle and noxious heat applied to the tail were reduced by 47%, 63%, 87% and 63%, respectively. The post-effects were also reduced both in terms of magnitude and duration. These results strongly suggest that serotonergic pathways partially involved in DNIC. They are discussed with reference to the descending control systems, originating from the caudal raphé, which modulate the transmission and/or the integration of nociceptive messages at the spinal level. The possible involvement of DNIC and 5-HT mechanisms to the hypo-algesic phenomena induced by hyper-stimulation is also suggested.

A method is described for the preparation of [carbonyl-11C]6-fluoromelatonin for intravenous injection and potential study of the melatonin system in vivo by positron emission tomography. The preparation is based on the acetylation of 6-fluoro-5-methoxytryptamine with NCA [1-11C]acetyl chloride (itself prepared from cyclotron-produced [11C]carbon dioxide) and purification by HPLC. It gives chemically and radiochemically pure [carbonyl-11C]6-fluoromelatonin in 35% radiochemical yield (from [11C]CO2, decay-corrected) within 35 min from the end of radionuclide production and with high specific activity e.g. 1.6 GBq/mumol (43 mCi/mumol) at the end of synthesis from 1.1 GBq (30 mCi) of [11C]carbon dioxide.

The effects of the serotonin precursor 5-HTP, were determined in a behavioral DNIC paradigm (increase in vocalization threshold after intraperitoneal injection of the algogenic agent, phenylbenzoquinone). This counter-irritation phenomenon was strongly potentiated by 5-HTP, such potentiation being blocked by the 5-HT receptor blocker, cinanserin. These results are in keeping with those of our recent single unit work in dorsal horn convergent neurons.

(1) Sixty-eight convergent dorsal horn neurones have been recorded at the lumbar level in anaesthetized intact rats. All cells received prominent A alpha and C fibre afferents and correspondingly could be activated by high and low threshold stimuli applied to the peripheral excitatory receptive field. (2) The activity of 67/68 of these neurones was powerfully inhibited by noxious stimuli applied to various parts of the body. Since non-noxious stimuli were ineffective in this respect, the term "diffuse noxious inhibitory controls" (DNIC) is proposed. (3) DNIC could be evoked by noxious pinch applied to the tail, the contralateral hind paw, the forepaws, the ears and the muzzle; the most effective areas were the tail and muzzle. Noxious heat applied to and transcutaneous electrical stimulation of the tail were extemely effective in eliciting DNIC as was the intraperitoneal injection of bradykinin. (4) DNIC strongly depressed by 60-100% both the C fibre response following suprathreshold transcutaneous electrical stimulation and the responses to noxious radiant heat. (5) The spontaneous activity and the responses to low threshold afferents induced either by A alpha threshold electrical or natural stimulation were also powerfully inhibited. (6) In the majority of cases, long lasting post-effects directly related to the duration of conditioning painful stimulus were observed.

1 Recordings were made from 18 neurones in the dorsal horn of the rat, anaesthetized with halothane. All cells received A- and C-fibre inputs and responded to innocuous and noxious stimuli applied to their excitatory receptive fields located on the extremity of the ipsilateral hindpaw. Transcutaneous application of suprathreshold (mean 3.2 T) 2 ms square-wave pulses to the centre of the receptive fields resulted in responses to A- and C-fibre activation being observed; a mean 32.4 +/- 6.0 C-fibre latency spikes were evoked per stimulus. 2 A high dose (20 micrograms) of morphine in 0.5 microliter sterile saline, microinjected into the periaqueductal grey matter (PAG) had no effect on the C-fibre-evoked activity of thirteen cells (72%) and facilitated 5 neurones (28%). Microinjection sites covered most of the PAG particularly the caudal medioventral zone. 3 A relatively high dose (6 mg kg-1, i.v.) of systemic morphine chloride, sufficient to elicit the direct spinal action of the opiate, inhibited all 5 cells tested. 4 We conclude that there is little evidence that the supraspinal action of morphine includes increased descending controls and depression of dorsal horn neurones in the rat.

(1) The effects of morphine upon the transmission of nociceptive messages at the spinal level have been investigated in the spinal rat. The responses of dorsal horn cells induced by the activation of C fibres were depressed in all cases in a dose-dependent fashion, this effect being reversed by the opiate antagonist naloxone. An estimation of the ED50 at the cellular level leads to the value of 6.3 mg/kg. The responses to A delta fibres were also depressed dose-dependently whereas the responses to A alpha fibres were unaffected. This is a confirmation in the rat of the differential effects of morphine on responses of convergent units elicited by the stimulation of different fibres, as previously described in the cat. (2) The hypothesis of the participation of serotonergic terminals in these effects has been checked by comparing the preceding results to those obtained in pCPA pretreated animals. Two populations of units were observed in the latter group: two-thirds of cells showed a dose-response curve similar to that of the non-pretreated group whereas the remaining one-third were unaffected either by morphine or naloxone. It is concluded that, at least, two mechanisms are involved in the depressive effects of morphine at the spinal level, serotonergic terminals being implicated in one of these. (3) The lowering of spinal cord serotonin content was associated with a decrease of both the size of the excitatory receptive field (34%) and the activities related to C fibre input (36%) of the recorded dorsal horn cells. This result is discussed with reference to the excitatory or sensitizatory effect of serotonin upon chemoreceptors related to pain.