Pain and child: a translational hypothesis on the pathophysiology of a mild type-2 diabetes model.

Pediatric pain management underwent many changes since the undertreatment of pain in children was reported in the literature in 1980. Increasing data also suggest that long-term behavioural effects can be observed in children, following pain episodes as early as in the neonatal period. Therefore, the knowledge about safe and effective management of pain in children should be applied with greater effectiveness into clinical practice. Other advances in the field include the findings of long-term residual behavioural and metabolic effects induced by pain experienced during the critical periods of development in laboratory animals. Recent data in laboratory animals and clinical data in children suggest that early repeated and/or severe pain and other stressful procedures applied in the perinatal periods may produce not only behavioral, but also important hormonal, immune and metabolic long-term effects. In this paper we shall report data on some metabolic conditions described in adult humans following disruption of hormonal-metabolic programming produced in the peri-natal period. Quite similar signs can be found between animal models and human conditions, most of them being connected with hypothalamus-pituitary-adrenal hormones (HPA) dysfunction. In addition, some signs in animal models, such as overweight and abdominal overweight are prevented by treatment with the µ- and δ-opioid receptor antagonist naloxone during the lactating period. This indicates that some long-term consequences following stress received during the early phases of life in mammals may be bound to the HPA system dysregulation, whereas others are bound to different (e,g., opioid) endogenous brain receptors and/or neuromediators alteration.

Formation of an adduct by clenbuterol, a beta-adrenoceptor agonist drug, and serum albumin in human saliva at the acidic pH of the stomach: evidence for an aryl radical-based process.

Clenbuterol (CLB) is an antiasthmatic drug used also illegally as a lean muscle mass enhancer in both humans and animals. CLB and amine-related drugs in general are nitrosatable, thus raising concerns regarding possible genotoxic/carcinogenic activity. Oral administration of CLB raises the issue of its possible transformation by salivary nitrite at the acidic pH of gastric juice. In acidic human saliva CLB was rapidly transformed to the CLB arenediazonium ion. This suggests a reaction of CLB with salivary nitrite, as confirmed in aerobic HNO(2) solution by a drastic decrease in nitric oxide, nitrite, and nitrate. In human saliva, both glutathione and ascorbic acid were able to inhibit CLB arenediazonium formation and to react with preformed CLB arenediazonium. The effect of ascorbic acid is particularly pertinent because this vitamin is actively concentrated within the gastric juice. EPR spin trapping experiments showed that preformed CLB arenediazonium ion was reduced to the aryl radical by ascorbic acid, glutathione, and serum albumin, the major protein of saliva. As demonstrated by anti-CLB antibodies and MS, the CLB-albumin interaction leads to the formation of a covalent drug-protein adduct, with a preference for Tyr-rich regions. This study highlights the possible hazards associated with the use/abuse of this drug.

Taurine administration during lactation modifies hippocampal CA1 neurotransmission and behavioural programming in adult male mice.

Taurine plays a role in neuronal development. In this study, we examined whether postnatal taurine administration influences the long-term consequences induced by mild neonatal stressors (10 min maternal deprivation plus sham injection, applied daily to neonatal mice up to 21 days). At 30 days of age stressed mice showed higher pain threshold both in the tail-flick--which measures mostly the spinal mechanisms of pain--and in the hot-plate test--which reflects mainly the supraspinal mechanisms of pain. The latter effect was prevented completely by neonatal taurine administration, while the tail-flick test was not affected, thus suggesting that spinal pain is not sensitive to taurine treatment. At 140 days of age, mice which were stressed during the neonatal period showed consistent decrease in immobility time in forced swimming test, and taurine did not influence this parameter. At the same age, the fear/anxiety axis, measured with elevated plus maze test, did not show any consistent changes. Electrophysiological experiments in brain slices obtained from adult mice showed that input-output curves in hippocampal CA1 were increased by taurine administration in lactation. Hence, neonatal administration of taurine might permanently modify the functioning of hippocampus, a brain area which is known to be crucial for learning and memory.

Nitric oxide is involved in the expression of neocortical spike-and-wave spindling episodes in DBA/2J mice.

This study investigated the possible role of nitric oxide (NO) in the development of neocortical spike-and-wave spindling episodes (S&W) of DBA/2J mice. The administration of distilled water did not modify either the number or duration of S&W in DBA/2J mice during the whole recording period (240 min). L-N(G)-nitro arginine methyl ester (L-NAME) (3-300 microg/mouse, i.c.v.) dose-dependently reduced the S&W of DBA/2J mice. This effect appeared 30 min after drug administration and lasted for the duration of the recording period (240 min). In addition, L-NAME treatment did not induce significant alterations of stereotyped behaviour such as licking, sniffing, chewing or tremors of the head and body and behavioural excitability, whereas the electroencephalogram desynchronized pattern was also significantly reduced. By contrast D-N(G)-nitro arginine methyl ester at the same doses did not affect S&W of mice. The inhibitory effect of L-NAME on S&W of mice was dose-dependently reversed by L-arginine (L-ARG, 3-300 microg/mouse, i.c.v.) but not by D-arginine. Finally, glyceryl trinitrate on its own (3-300 microg/mouse, i.c.v.) significantly increased the S&W of mice and it was also able to reverse the inhibition on S&W of mice operated by L-NAME. These results provide evidence that NO may play a significant role in the development of brain excitability.

ALTEA: anomalous long term effects in astronauts. A probe on the influence of cosmic radiation and microgravity on the central nervous system during long flights.

The ALTEA project participates to the quest for increasing the safety of manned space flights. It addresses the problems related to possible functional damage to neural cells and circuits due to particle radiation in space environment. Specifically it aims at studying the functionality of the astronauts' Central Nervous Systems (CNS) during long space flights and relating it to the peculiar environments in space, with a particular focus on the particle flux impinging in the head. The project is a large international and multidisciplinary collaboration. Competences in particle physics, neurophysiology, psychophysiology, electronics, space environment, data analyses will work together to construct the fully integrated vision electrophysiology and particle analyser system which is the core device of the project: an helmet-shaped multi-sensor device that will measure concurrently the dynamics of the functional status of the visual system and passage of each particle through the brain within a pre-determined energy window. ALTEA is scheduled to fly in the International Space Station in late 2002. One part of the multi-sensor device, one of the advanced silicon telescopes, will be launched in the ISS in early 2002 and serve as test for the final device and as discriminating dosimeter for the particle fluences within the ISS.

Effects of cysteamine on nociception in mice.

AIM: The present study was carried to study the effects of cysteamine on nociception in mice. METHODS: The pain assays were the hot plate and the tail flick test. RESULTS: When cysteamine, a drug well known as a somatostatin depletor, was administered 1 and 4 but not 24 h before the tests (hot plate, tail flick), the nociceptive threshold was elevated when the drug was administered at high doses (50 and 100 mg/kg) while at a lower dose (10 mg/kg), it was able to elevate the nociceptive threshold in the hot plate test only. In the hot plate as well the tail flick test cysteamine effects are reversed by naloxone administration and potentiated by morphine administration, whereas neither somatostatin nor cyclo-(7-aminoheptanoyl-Phe-D-Trp-Lys-Thr[Bzl], a reported somatostatin antagonist, changes cysteamine effects. CONCLUSION: These results suggest that cysteamine effects on the nociceptive threshold in the hot plate and tail flick test may be mediated by cysteamine interference with the opioid system.

Purinoreceptors are involved in the control of acute morphine withdrawal.

The effects exerted by P1 and P2 purinoceptor agonists and antagonists on the acute opiate withdrawal induced by morphine were investigated in vitro. Following a 4 min in vitro exposure to morphine, the guinea-pig isolated ileum exhibited a strong contracture after the addition of naloxone. The P1 purinoceptor agonist, adenosine, was able dose-dependently to reduce morphine withdrawal whereas alpha,beta-methylene ATP (APCPP), a P2 purinoceptor agonist, increased morphine withdrawal. Caffeine, a P1 purinoceptor antagonist, was able significantly and in a concentration dependent manner to increase morphine withdrawal whereas quinidine, a P2 receptor antagonist, reduced it. The results of our experiments indicate that both P1 and P2 purinoceptor agonists and antagonists are able to influence opiate withdrawal in vitro, suggesting an important functional interaction between the purinergic system and opioid withdrawal.

Arachidonic acid and its metabolites are involved in the expression of neocortical spike-and-wave spindling episodes in DBA/2J mice.

This work was undertaken to study the effects of dexamethasone, indometacin and mifepristone plus dexamethasone on the neocortical spike-and-wave spindling episodes (S&W) in the electrocorticogram of DBA/2J mice. Our data indicate that both dexamethasone and indometacin (1, 10, 100 microg kg(-1), i.p.) reduced the S&W of DBA/2J mice. This effect appeared 30 min after drug administration and lasted for the duration of the recording period (240 min). Mifepristone, a glucocorticoid receptor antagonist (1, 10, 100 microg kg(-1), i.p.), injected 2 h before dexamethasone, totally blocked the steroid effect. These results indicate that both dexamethasone and indometacin significantly reduce the S&W of DBA/2J mice, suggesting a possible involvement of arachidonic acid and its metabolites in the development of brain excitability.

Dexamethasone blocking effects on mu- and delta-opioid-induced seizures involves kappa-opioid activity in the rabbit.

Previous data indicate that intracerebroventricular administration of agonists for mu- and delta-opioid receptors induces limbic seizures in rats, but no data are reported in rabbits. We found that the mu- and delta-opioid peptides [D-Ala(2)-N,Me-Phe(4)-Gly(5)-ol]enkephalin (DAMGO), beta-endorphin and deltorphin II, induced EEG non-convulsive hippocampal seizures, and changes in hippocampal background EEG, physical parameters and overt behaviour after central administration. Dexamethasone pre-treatment prevented DAMGO-, deltorphin II- and beta-endorphin-induced seizures as well as changes in background EEG, physical parameters and overt behaviour induced by mu-opioid agonists. Dexamethasone antagonism on opioid action was blocked by pre-treatment with a protein synthesis inhibitor, cycloheximide or by the kappa-opioid antagonist nor-binaltorphimine. Our data suggest that dexamethasone influences opioid actions at mu- and delta-receptors via a protein synthesis mechanism involving kappa-opioid receptors.

Clonidine-induced antinociception and locomotor hypoactivity are reduced by dexamethasone in mice.

The effects of dexamethasone pretreatment on clonidine-induced antinociception and locomotor hypoactivity were investigated in mice. In the hot-plate and the tail-flick tests, dexamethasone administered intraperitoneally at a dose of 1 mg kg(-1), 30 or 60 min before clonidine, reduced clonidine antinociception in both tests and reduced clonidine-induced locomotor hypoactivity in the activity cage. When administered 15 min before clonidine, dexamethasone had no effect on clonidine antinociception. A higher dexamethasone dose (10 mg kg(-1)) induced the same effects observed at a dose of 1 mg kg(-1) in the hot-plate and the tail-flick tests, but the former dose had a stronger effect on locomotor hypoactivity. Dexamethasone (10 ng/mouse) administered intracerebroventricularly 30 min before clonidine was also able to reduce both clonidine-induced antinociception and locomotor hypoactivity. The protein synthesis inhibitor, cycloheximide, administered intraperitoneally at the dose of 10 mg kg(-1), 2 h before clonidine, was able to prevent dexamethasone effects on clonidine-induced antinociception. The glucocorticoid receptor antagonist RU-38486, administered intracerebroventricularly at the dose of 1 ng/mouse, was also able to block dexamethasone effects on clonidine-induced antinociception and locomotor hypoactivity, whereas both cycloheximide and RU-38486 per se did not influence pain sensitivity or locomotor activity. These results suggest that the dexamethasone effects on clonidine-induced antinociception and locomotor hypoactivity depend on the stimulating effects that dexamethasone exert, on the protein synthesis via the glucocorticoid receptor in the brain.

The ALTEA facility on the International Space Station.

The ALTEA project studies the problems related to possible functional damage to the Central Nervous System (CNS) due to particle radiation in space environment. The project is a large international and multi-disciplinary collaboration. The ALTEA instrumentation is an helmet-shaped multi-sensor device that will measure concurrently the dynamics of the functional status of the visual system and the passage of each particle through the brain within a pre-determined energy window. ALTEA is scheduled to fly in the International Space Station in February 2003. One part of the multi-sensor device, one of the advanced silicon telescopes, will be launched in the ISS in early 2002 and serve as test for the final device and as discriminating dosimeter for the particle fluences within the ISS.

Clinical and pharmacological profile in a clenbuterol epidemic poisoning of contaminated beef meat in Italy.

Long-acting beta adrenergic agonists, such as clenbuterol accumulate in the liver, but not meat of treated farm animals, and result in epidemic poisonings in consumers. We describe an outbreak of poisoning in 15 people, following the consumption of meat. Clinical symptoms (distal tremors, palpitations, headache, tachipnoea-dyspnoea, and also moderate hyperglycaemia, hypokalemia and leucocytosis) were seen in nine hospitalised patients, starting about 0.5-3 h after poisoning, and disappearing within 3-5 days later. Clenbuterol was found in the urine of all the symptomatic patients, at higher levels than pharmacokinetic computing (mean level 28 ng/ml, 36 h after ingestion), based on the levels found in the meat (1140-1480 ng/g edible tissue). Thus, epidemic poisoning can be produced following the consumption of contaminated meat. The need for a better definition of pharmaco- and toxico-kinetics, not only for drugs ingested as parent drug, but also when ingested as residues with animal tissues, is recommended.

Guinea-pigs treated with beta-endorphin fragments DTgammaE and DEgammaE exhibit reduced morphine inhibition of electrically-induced contractions.

The effect exerted by two gamma-endorphin derivatives (DTgammaE and DEgammaE) was investigated on morphine-induced inhibition on the electrically contractions of guinea pig ileum in vitro. Morphine (1x10(-8)-5x10(-8)-1x10(-7) M) dose dependently and significantly reduced the E.C. of guinea pig ileum, IC50=6.5x10(-8) M (Confidence limits: 3.7x10(-8)-9.1x10(-8)). DTgammaE and DEgammaEper se (1x10(-6)-5x10(-6)-1x10(-5) M) did not modify significantly the E.C. of guinea pig ileum. Furthermore, DTgammaE or DEgammaE injection 10-30-60 min before morphine, did not affect the inhibitory effect of morphine on the E.C. of guinea pig ileum. By contrast, ilea from guinea-pigs treated for 4 days with DTgammaE or DEgammaE (1 mg/Kg/i.p.) resulted less sensitive to the inhibitory effect of morphine, IC50=8.3x10(-7) M (Confidence limits: 1.4x10(-6)-3.5x10(-7)) for DTgammaE and IC50=7.7x10(-7) M (Confidence limits: 2.7x10(-6)-8.7x10(-7)) for DEgammaE. Our results indicate that chronic treatment of guinea pigs with DTgammaE or DEgammaE induces a significant reduction of the inhibitory effect of morphine on the E.C. of guinea-pig ileum thus confirming an important functional interaction between gamma-endorphin derivatives and opioid system.

Postnatal Stress in Mice: Effects on Body Fat, Plasma Lipids, Glucose and Insulin.

Mice pups were exposed to stressful stimuli everyday during the first 3 weeks of life. Body weight, food intake and spontaneous locomotor activity, triglycerides, cholesterol, phospholipids, glucose and insulin basal levels, as well as epididymal fat pad weight and its cell volume were measured in stressed and control animals. Results indicated that postnatal stressful manipulations induced an increase in body weight, epididymal fat pad weight and its cell volume, as well as in insulin, glucose, cholesterol and triglycerides plasma levels, at 4 months of age. No significant changes in food consumption, locomotor activity and phospholipids plasma levels were found. Present data suggest that early stressful manipulations may induce residual effects on lipid and glucid metabolism.

Orphanin FQ reduces morphine-induced dopamine release in the nucleus accumbens: a microdialysis study in rats.

The effects induced by orphanin FQ (OFQ) on morphine-induced dopamine (DA), 3,4-dihydroxyphenilacetic acid (DOPAC) and homovanillic acid (HVA) release in the nucleus accumbens were studied in rats by using microdialysis with electrochemical detection. Morphine administered intraperitoneally (i.p., 2, 5 and 10 mg/kg) dose-dependently increased DA and metabolites release in the nucleus accumbens. OFQ intracerebroventricularly (i.c.v.) administered at doses of 2, 5 and 10 nmol did not change DA and metabolites release in the nucleus accumbens. OFQ (10 nmol) administered i.c.v. 15 min before morphine (5 and 10 mg/kg, i.p.) significantly reduced morphine-induced DA and metabolites release in the nucleus accumbens. These effects suggest that OFQ may regulate the stimulant action linked to morphine-induced DA release in the nucleus accumbens.

Effects of cholinergic drugs on neocortical EEG and flash-visual evoked potentials in the mouse.

The effects of single intraperitoneal injection of two cholinesterase inhibitors, physostigmine (PHY; 0.01, 0.025, 0.05, 0. 1, 0.2 mg/kg) and heptylphysostigmine (HEP; 0.5, 2, 6 mg/kg) on electroencephalographic (EEG) activity and flash visual evoked potentials (f-VEP) in the occipital cortex were compared in DBA/2 mice. EEG spectral analysis of awake periods showed that PHY at all doses and HEP at 2 mg/kg induced an increase of power in the 4.25- to 7-Hz frequency band. Furthermore, PHY at the higher doses and HEP at all doses induced a decrease of power in the 7.25- to 12-Hz frequency band, while the lower doses of PHY (0.01, 0.025 mg/kg) produced an increase of this band. EEG effects elicited by the two drugs were similar, when doses displaying analogous biochemical effects (acetylcholinesterase inhibition) were used (i.e. 0.01 and 0. 025 mg/kg of PHY versus 0.5 and 2 mg/kg of HEP). PHY and HEP induced similar changes in f-VEPs. Amplitudes of early and late components (P1N1, N1P2, P4N4 and particularly N1P3) were enhanced, while amplitudes of middle components were depressed after all doses. The peak latency measures were generally delayed, even though, after the lower doses, a trend to a latency reduction was evident in late components. This finding might indicate a possible effect on stimulus speed diffusion by 'low therapeutic' doses, analogous to the ones used in men. Our data show that both drugs are effective in modifying EEG and f-VEP parameters connected with brain cholinergic function, although in a very narrow dose range.

Adrenocorticotropin-related modulation of the human EEG and individual variability.

During a 6-h period in resting conditions, the blood concentrations at rest of cortisol, glucose and the adrenocorticotropic hormone (ACTH) varied spontaneously within physiological ranges in eight healthy male volunteers (24.5+/-1.7 years), without pulsatile changes, correlation among variables, or indications of stress response. The power of the 6.5-14.0 Hz physiological 'alpha' rhythm of the electroencephalogram (EEG) proved inverted-U correlated with the ACTH concentration (with maximum power at 12-14 pmol/l ACTH) but was independent from the extent of ACTH change or from cortisol/glucose concentrations. Two subgroups of subjects with low/high EEG power values could be separated depending on ACTH concentration, with estimated cut-off at 7-8 pmol/l. A direct ACTH modulation of brain electrophysiology or common factors (e.g. the corticotropin-releasing hormone) pacing both ACTH and EEG are suggested and may account for individual EEG differences.

Effects of neonatal treatment with Tyr-MIF-1 and naloxone on the long-term body weight gain induced by repeated postnatal stressful stimuli.

Stressful stimuli repeatedly applied during the first postnatal weeks can induce body weight gain in the mouse during adulthood. This effect can be prevented by injecting naloxone concomitantly with stress. The peptides belonging to the Tyr-MIF-1 family have a great modulating activity on numerous stress-induced phenomena. The aim of the present work was to compare the effect of repeated neonatal injections of Tyr-MIF-1 or naloxone on the long-term body weight gain induced by a stressing procedure applied daily during the first three weeks of life. The results indicate that although naloxone blocked the development of the stress-induced effects, Tyr-MIF-1 potentiated them.

Clenbuterol residues in non-liver containing meat as a cause of collective food poisoning.

beta 2-adrenergic agonists, particularly clenbuterol, are illegally used as growth promoters to obtain lean in meat. Their administration in feedlots can constitute a severe risk for animal welfare and exposes consumers to involuntary drug consumption at pharmacological active concentrations. Reported poisoning episodes have been associated with the consumption of beef liver where clenbuterol residues concentrate. In August 1996, 62 persons asked for medical help at the emergency rooms of 2 hospitals near the city of Caserta (Italy). Their clinical profile was characteristic of previously occurring clenbuterol intoxication, which reported superventricular extrasystoles and atrial fibrillation. All patients had non-liver beef meat consumption 10-30 min to 2-3 h before symptoms developed. An ELISA screening test specific for clenbuterol confirmed the drug's presence. Definitive confirmation of clenbuterol and determination of the drug content in meat samples were obtained by GC-MS, using 2 different derivatization. Concentrations in the meats ranged from 0.8 to 7.4 mg/kg. These analytical data provided evidence of the seriousness of the poisoning and helped the National Health System identify other possible misinterpreted cases. This case demonstrates that clenbuterol poisoning can also occur after consumption of beef meat other than liver.