The site of action of naloxone in suppressing food and water intake in rats

Previous studies have shown that naloxone causes a decrease in food and water intake; however, the site of this action has not been determined. We investigated this problem by giving bilateral injections of 15 μg/rat of naloxone into the lateral ventricles of cannulated, food and water deprived rats. This treatment caused a significant decrease in food intake when compared to saline injected controls. Water intake in naloxone-treated animals did not differ significantly from that of saline-treated controls during the one hour test period. The total dose of naloxone given centrally, 15 μg, did not produce a change in eating or drinking if given peripherally. The findings imply that naloxone exerts its effect on food intake at a central site. A dose-related and significant suppression of water intake was seen after treatment with nalaxone peripherally (1, 3, and 10 mg/kg, i.p.) in rats with either subdiaphragmatic vagotomy (vag) or a sham vagotomy (sham). Although a significant suppression of food intake was seen in the sham rats, no supression of food intake was seeen in the vag rats at any dose of naloxone tested. In rats pretreated with methyl atropine (5 mg/kg, i.p.), naloxone (3 mg/kg, i.p.) was equivalent to saline in that it did not decrease food intake. However, nalaxone did cause a significant decrease in water intake in methylatropine pre-treated rats. These results suggest that the suppression of food intake by naloxone has a central site of action which is mediated by the vagus, and specifically by vagal efferents, since the effect was blocked by methylatropine. The results also suggest that naloxone's effect on water intake is mediated by a different mechanism than that involved with food intake.     

Long-term enhancement of morphine hypalgesia as a result of periodic blockade of opiate receptors using naloxone

A significant enhancement of the analgetic effect of morphine (6 mg/kg, subcutaneously; tail withdrawal reflex at 60 degrees C) was observed in rats 3-4 hours after single naloxone (1 mg/kg) administration. Periodical naloxone injection (0.5 mg/kg, subcutaneously, 3 times per day at 3.5-hour intervals for 3 days) led to a prominent and long-term (testing on the 20th and 105th hour after the last naloxone administration) enhancement of morphine analgesia (2.6 mg/kg subcutaneously) and insignificant inhibition of stress analgesia during two-hour immobilization of animals. These modifications of morphine and stress analgetic effects are considered a result of adaptive changes of opiate receptors after their blockade.     

Changes in total and free tryptophan levels in serum following acute morphine administration in arthritic rats

In ‘arthritic’ rats a decrease in total tryptophan and an increase in free tryptophan levels was observed in serum after morphine administration (10 mg kg, s.c.). These changes were maximum within 15 and 30 min after injection.A decrease in total and an increase in free tryptophan levels in serum were observed 30 min after naloxone administration (1 mg/kg, i.m.).An increase in tryptophan and 5-hydroxyindoleacetic acid levels was also observed in the brain after morphine and naloxone.These observations suggest that the rise in 5-hydroxytryptamine synthesis provoked by morphine may be partly related to an increase in the availability of tryptophan from blood. However, the analgesia induced by the opiate appears unlikely to be directly related to this effect.     

The effect of naloxone on the activation, inducible by chronic alcoholic intoxication, of the membrane-bound form of enkephalin convertase in the midbrain and hypothalamus of rats]

The activity of membrane-bound and soluble enkephalin convertase was determined with dansyl-Phe-Leu-Arg as substrate in midbrain, including hypothalamus, of Wistar rats, who were given ethanol (20% solution i.g., 9-15 g/kg per day during 4 days) or naloxone (2 mg/kg i.p. twice a day during 4 days) or their combination. It was shown that activation of membrane-bound enzyme, observed after alcohol treatment of rats, didn't develop by combined ethanol-naloxone administration. It's supposed that alcohol-stimulating effect on this enzyme realizes throughout the hyperstimulation of opioid receptors by enkephalins and, possible, by other opioid-active compounds.     

Strain differences in the dose-response curves of adrenalectomized, starved-refed rats to dehydroepiandrosterone (DHEA)

The effects of adrenalectomy and dehydroepiandrosterone (DHEA) doses (0, 15, 30, 60, 120 and 240 mg/kg/day ip) on hepatic enzyme activity and lipid content and on the amount of epididymal fat pad lipid were studied in starved-refed BHE and Sprague-Dawley rats. BHE rats had significantly greater relative liver size, glucose-6-phosphate dehydrogenase (G6PD) and malic enzyme (ME) activities, and percentage liver lipid but less epididymal fat pad lipid than Sprague-Dawley rats. Adrenalectomized (ADX) rats consumed significantly less food, gained less weight per day, and had less lipid in their livers and fat pads than intact rats. As the level of DHEA increased from 0 to 240 mg/kg/day there was a significant linear decrease in average daily weight gain, food intake, G6PD activity, and percentage liver lipid. At the 15 mg/kg/day dose, G6PD activity was significantly reduced without reductions in the other parameters measured. At the 120 mg/kg/day dose, however, weight gain, food intake, G6PD activity, and percentage liver lipid were significantly lower than that of the controls. At this dose DHEA treatment reduced food intake by 17% whereas it diminished average daily weight gain and G6PD activity by 30 and 56%, respectively. The 240 mg/kg/day dose of DHEA significantly reduced food intake, weight gain, liver lipid, G6PD activity, and ME activity. Intact and ADX BHE rats reduced their G6PD activity and liver lipid more rapidly than Sprague-Dawley rats as the level of DHEA administered increased. ADX Sprague-Dawley rats receiving DHEA had greater liver lipid content and enzyme activity than their intact counterparts whereas the reverse situation was true in BHE rats. These data indicate that the effect of DHEA on body weight gain, food intake, and hepatic and peripheral adiposity are dependent on the strain of rat, the adrenal status, and the DHEA dose.     

Naloxone effect on the neurological deficit induced by forebrain ischemia in rats

The effect of naloxone upon neurologic deficit was evaluated in a model of transient forebrain ischemia in rats. Awake male Wistar rats were subjected to a 30 minute ischemia by occluding both common carotid arteries 8 days after cauterizing vertebral arteries. Administration of naloxone 1 or 5 mg/kg iv 10 minutes after carotid occlusion or 1 mg/kg iv one hour after clamp removal failed to reduce immediate and tardive neurologic postischemic deficits. On the other hand, in rats treated by a dose of 1 mg/kg naloxone 10 minutes after carotid occlusion and perfused with an additional dose of 2 mg/kg/h for 80 minutes, neurologic score was improved one hour after ischemia. However mortality was not decreased whatever was the modality of naloxone administration. This result confirms previous data showing that naloxone exerts a protective effect when given at sufficiently high dosage.     

D-amphetamine-induced hypothermia and hypermotility in rats: Changes after systemic administration of beta-endorphin

Systemically administered beta-endorphin was tested in rats for its ability to modify the hypothermia and hypermotility induced by d-amphetamine. Colonic temperature and motor activity were measured in a cold (4°C) ambient temperature in animals given IP injections of beta-endorphin (0.1, 1.0, or 3.0 mg/kg), naloxone (10 mg/kg), or morphine (30 mg/kg). The same measurements were taken in animals given beta-endorphin (1.0 mg/kg) in combination with naloxone or saline pretreatment and d-amphetamine (15 mg/kg) or saline post-treatment. Morphine alone had a biphasic effect on thermoregulation, but did not affect d-amphetamine-induced hypothermia. Activity scores were decreased by morphine, in both d-amphetamine and saline treated animals. The thermal response of rats to beta-endorphin alone was variable, depending on dosage, but all 3 dosages partially blocked the hypothermic effect of d-amphetamine. Naloxone blocked the thermal effects of both beta-endorphin and d-amphetamine. Motor activity tended to be decreased by naloxone, regardless of amphetamine treatment, but beta-endorphin tended to increase activity in amphetamine-treated animals and reduce it in saline-treated controls. In their actions on both thermoregulation and activity, naloxone and beta-endorphin appeared to interact independently with d-amphetamine, often producing effects in the same direction, but in combination, they tended to be mutually inhibitory.     

Evidence for sedative effects of low doses of morphine in mice involving receptors insensitive to naloxone

Low doses of morphine (0.30–2.5 mg/kg) decrease in a dose-dependent manner spontaneous climbing behaviour in mice. This effect is not modified by administration of naloxone at doses up to 1.25 mg/kg. These morphine doses do not modify the locomotor activity but, when they are associated with naloxone (0.5 mg/kg), an obvious inhibition occurs. In rats, a hyperactivity follows the akinesia produced by a morphine administration (10 mg/kg). This hyperactivity is changed into a significant hypokinesia when the animals are treated with naloxone (0.05 mg/kg). These results might reveal a dual effect of low doses of morphine, the excitatory effect of morphine being antagonized by naloxone whereas no action on the sedative effect is observed.     

Effects of Opiates on High-Affinity Ca2+,Mg2+-ATPase in Brain Membrane Subfractions

The effect of a single administration of morphine sulfate (15 mg/kg, s.c. or 30 mg/kg, i.p., 30 min) on Ca2+-stimulated Mg2+-dependent ATPase activity was investigated in synaptosomal plasma membranes (SPM) prepared from rat cortex. Morphine produced a significant decrease in Ca2+,Mg2+-ATPase activity in synaptosomal fractions (SPM 1 + 2) known to contain a high density of opiate receptors and calmodulin-dependent Ca2+,Mg2+-ATPase. However, in another subpopulation (SPM 3) that contains fewer opiate receptors and less enzyme activity, no such decrease in the enzyme activity was observed after the opiate administration. The decrease in Ca2+,Mg2+-ATPase activity seen in SPM 1 + 2 was specifically antagonized by the opiate antagonist naloxone hydrochloride (2 mg/kg, s.c.) when given 15 min before morphine administration. Mg2+-ATPase was not altered either by morphine or by a naloxone-morphine combination. These findings give further evidence for the role of intracellular Ca2+ in mediating many of the acute effects of opiates.     

In vivo radioprotection by alpha-TMG: preliminary studies

alpha-TMG is a novel water-soluble derivative of Vitamin E that has shown excellent antioxidant activity. The parent compound has demonstrated protection against radiation induced chromosomal damage in vivo. Hence, the preliminary experiments to determine the radioprotective activity of alpha-TMG were carried out in adult Swiss albino mice. Acute toxicity of the drug was studied taking 24h, 72 h and 30 day mortality after a single intraperitoneal injection of 500-2000 mg/kg body weight of the drug. The drug LD(50) for 24h and 72 h/30 day survival were found to be 1120 and 1000 mg/kg body weight, respectively. The optimum time of drug administration and drug dose-dependent effect on in vivo radiation protection of bone marrow chromosomes was studied in mice. Injection of 600 mg/kg of the drug 15 min before or within 5, 15 or 30min after 3Gy whole body gamma radiation resulted in a significant decrease in the aberrant metaphases percent at 24h post-irradiation; the maximum effect was seen when the drug was given immediately after irradiation. Injection of 200-800 mg/kg TMG within 5 min of irradiation with 3 Gy produced a significant dose-dependent reduction in the radiation induced percent aberrant metaphases and in the frequency of micronucleated erythrocytes at 24h after exposure, with a corresponding decrease in the different types of aberrations. The optimum dose for protection without drug toxicity was 600 mg/kg body weight. At this dose, TMG produced 70 and >60% reduction in the radiation induced percent aberrant metaphases and micronucleated erythrocytes, respectively. The high water solubility and effectiveness when administered post-irradiation favor TMG as a likely candidate for protection in case of accidental exposures.     

Effects of morphine and naloxone administered to pregnant rats on the adrenal glands and testes of the offspring]

Relative testis and adrenal weights, testosterone and corticosterone levels in the blood were determined in 9-, 16-day and 2-month-old rats born to females injected with morphine (10 mg/kg/day), naloxone (10 mg/kg/day) or saline throughout the 15-18 days of gestation. Opioid receptors agonist morphine caused a long-lasting inhibition of the testes and activation of the adrenals. Saline injections to the females, that are known to be a stressor for them, also inhibited the testis of the neonatal offsprings. The block of the opioid receptors by the naloxone prevented the effect of prenatal stress on the testis, but inverted the negative correlation between the testes and the adrenals, that can be observed in the normal development.     

Influence of antioxidant effect of stobadine derivative in condition of kidney ischemia-reperfusion in a pre-clinical experiment (effect in prophylaxis)

The goal of the study was to monitor the antioxidative effect of stobadine derivative in the conditions of ischemia-reperfusion of laboratory rat kidney tissue. The animals were divided by random selection into 5 groups (n = 10). The treated groups were given stobadine derivate in peroral doses of 5, 10 and 20 mg/kg in 0.5 % solution of Avicel once a day; the placebo group was given only the solution of Avicel. The last group was an intact group (without ischemia-reperfusion and without treatment). After conclusion of medication on the 15th day all animals were subjected to kidney tissue ischemia (60 min.) followed by reperfusion (10 min.). All animals were subsequently exsanquined and single identification of superoxiddismutase, glutathion peroxidase, total antioxidative capacity, and malondialdehyde level in the blood were determined. Kidneys were recovered for histopathological examination. A statistically significant decrease of the superoxiddismutase and statistically significant increase of the glutathione peroxidase catalytic activity in the treated groups compared to the groups of placebo and intact was discovered. There was also a statistically highly significant increase of total antioxidative capacity in the treated groups compared to the groups of placebo and intact. A statistically significant decrease of malondialdehyde level was identified in the treated groups compared to the groups of placebo and intact. The results of biochemical examination show a protective antioxidative effect of stobadine derivative. The results of histopathological examination support this assumption.     

Effects of morphine and naloxone on glucose metabolism in uterine strips from ovariectomized and non-ovariectomized restricted diet rats

The effect of underfeeding over glucose metabolism in uteri isolated from ovariectomized and non-ovariectomized rats subjected to a restricted diet for 25 days (50% of the normal food intake), was studied. Underfeeding decreases (14)CO(2) formation from U(14) C-glucose in intact animal uteri. While in ovariectomized rats (25 days), the effect is the opposite. The addition of morphine 10(-6) M to the medium does not affect rats fed ad libitum. However, (14)CO(2) levels increase significantly in intact animals receiving a restricted diet. In ovariectomized rats morphine does not show any activity, regardless of the type of diet rats were subjected to. None of the rat groups seems to be sensitive to naloxone 10(-6) M. The s.c. injection of morphine (4 mg.kg (-1)) increases glucose metabolism only in intact rats provided with a restricted diet, while naloxone (2.5 mg.kg (-1) ) produces a decrease of ( 14)CO(2) in ovariectomized underfed animals. To conclude, morphine either 'in vivo' or 'in vitro' is active only in uteri from intact rats subjected to underfeeding. Naloxone produces a decrease in (14)CO(2) production, particularly when it is s.c. injected to ovariectomized rats undergoing a dietary restriction. Since the uterus does not react to naloxone, the effect of the opiod blocker may be the result of endogenous opioids originated in other tissues.     

Naloxone stimulation of luteinizing hormone release in prepubertal female rats; role of serotonergic system

The purpose of this study was to determine whether naloxone stimulated LH release via a serotonergic mechanism. Injection of naloxone hydrochloride (2 mg/kg B.W.) into 25-day old female prepubertal rats resulted in a significant elevation in serum LH 30 min later. Injection of this dose of naloxone together with morphine sulfate (2 or 5 mg/kg B.E.) resulted in inhibition of naloxone-induced LH release. When rats were first injected with 5-hydroxytryptophan (5-HTP) to increase hypothalamic serotonin content, naloxone failed to increase serum LH levels. On the other hand, when parachlorophenylalanine (PCPA) was given first to reduce hypothalamic serotonin content, naloxone-induced LH release was potentiated. Morphine failed to inhibit the naloxone-induced rise in serum LH when PCPA was first administered. Neither 5-HTP nor PCPA, when injected alone, altered serum LH values. These results suggest that naloxone promotes LH release by reducing hypothalamic serotonergic activity, and morphine inhibits LH release by increasing hypothalamic serotonergic activity. This does not exclude possible involvement of other neurotransmitters.     

Suppression of GABA-induced abstinence behaviour in naive rats by morphine and bicuculline.

Intraperitoneal administration of n-dipropylacetate (DPA) to naive rats produced abstinence behaviour including shaking, digging, hunchback posture, piloerection and ptosis during 15 min and increased motor activity considerably. Treatment with a subconvulsive dose of the GABA antagonist bicuculline suppressed this DPA-induced abstinence behaviour, indicating that GABA was increased at receptor sites. Also morphine in a low dose of 1 mg/kg suppressed this behaviour, while administration of naloxone after morphine treatment could release the abstinence behaviour. Simultaneous treatment with morphine and naloxone or naloxone alone were without effect. The administration to DPA treated rats of doses higher than 1 mg/kg morphine resulted in a severe depression of motor activity. It is concluded that an increased availability of GABA at its receptor sites plays an important role in the behaviour observed after DPA administration. The experiments with morphine and naloxone suggest that morphine receptors are involved in DPA-induced abstinence behaviour.     

Repeated dermal toxicity of technical HCH and methyl parathion (50EC) to female rats (Rattus norvigicus).

Repeated dermal application of hexachlorocyclohexane (HCH; 100 mg/kg/day) or methyl parathion (2 mg/kg/day) individually or in combination for 7, 15 and 30 days produced pathomorphological changes in skin, liver, kidney and brain of female rats along with significant enzymatic alterations in the activity of transaminase, alkaline phosphatase lactic dehydrogenase and acetylcholinesterase. The two insecticides in combination though produced severe toxicity on day 30 than at other periods, the changes were not suggestive of any additive or potentiation effect at the test doses.     

Involvement of vagal opioid receptors in respiratory effects of morphine in anaesthetized rats.

Respiratory effects of morphine injection to the femoral vein were investigated in urethane and chloralose anaesthetized and spontaneously breathing rats, prior to and after midcervical vagotomy. Bolus injection of morphine HCl at a dose of 2 mg/kg of body weight induced depression of ventilation in all rats, due to the significant decrease in tidal volume and to the decline in respiratory rate both pre- and post-vagotomy. Expiratory apnoea of mean duration of 10.0+/-3.4 s was present in the vagally intact rats only. Bilateral midcervical section of the vagus nerve precluded the occurrence of apnoea. Prolongation of the expiratory time (T(E morphine) / T(E control)), which amounted to 10.7+/-2.2-fold in the intact state, was apparently reduced to 1.5+/-0.3-fold after division of the vagi. Morphine significantly decreased mean arterial pressure (MAP) at 30 s after the challenge, the effect persisted for not less than 1 minute and was absent in vagotomized rats. The respiratory changes evoked by morphine reverted to the control level after intravenous injection of naloxone at a dose of 1 mg/kg. Results of this study indicate that opioid receptors on vagal afferents are responsible for the occurrence of apnoea and hypotension evoked by morphine.     

Decreased Histamine-Stimulated Phosphoinositide Hydrolysis in the Cerebral Cortex of a Rat Line Selectively Bred for High Alcohol Preference

This study sheds light on the comparative analysis of agonist-stimulated phosphoinositide (PI) hydrolysis in the cerebral cortex of alcohol-naive rats from established lines selectively bred for low alcohol preference (LAP) and high alcohol preference (HAP). The effect of histamine (1.0 mM), but neither norepinephrine (0.1 mM) nor carbachol (0.5 mM), on PI hydrolysis was significantly reduced in HAP rats (0.4 +/- 5.0 fmol/mg protein [3H]inositol phosphates formed over basal) compared with LAP rats (25.5 +/- 10.0 fmol/mg protein). The contents of monoamines (dopamine, norepinephrine, and serotonin) and histamine in the cerebral cortex did not significantly differ between LAP and HAP rats, nor did the contents of their metabolites, except 3-methoxy-4-hydroxyphenylglycol (one of the metabolites of norepinephrine) and N(tau)-methylhistamine, which was not detected in our system. The histamine stimulatory effect was unchanged in the cerebral cortex of an intact Wistar rat that was treated with intraperitoneal injection of alcohol (1.0 g/kg once per day for 14 days). The results of the current study indicate that the decrease in the histamine effect on PI hydrolysis in HAP rats might be attributed to that particular rat line.     

Effects of naloxone and cholecystokinin on food and water intake in vasopressin-deficient rats (Brattleboro strain)

Opioid peptides and cholecystokinin (CCK) have been shown to play a role in regulation of feeding behavior. Another neuropeptide that has recently been suggested to be involved in feeding is vasopressin. We explored possible interactions between opiates, CCK and vasopressin in feeding regulation by studying feeding suppression produced by naloxone and CCK in Brattleboro (DI) rats, which are homozygous for diabetes insipidus and lack the ability to synthesize vasopressin. Ten DI and 15 age-matched Long Evans (LE) rats were food deprived for 14 hours on two different days and then injected with naloxone (2.5 mg/kg) on one day or saline on the other. Thirty minutes later the food was returned and food and water consumption were measured after 1, 3 and 4 hr. Naloxone suppressed the food consumption of both DI and LE rats but the suppression was greater for the DI rats. This result was specific to feeding as water consumption was suppressed in LE more than in DI rats. Two weeks later, the same rats were food deprived for 6 hours on two different days and then injected with CCK-8 (2.5 micrograms/kg) on one day and with saline on the other. Food was returned one minute after the injection and food and water consumption were measured 30 and 60 minutes later. Food intake was reduced equally for both DI and LE rats. Water intake was not reduced. The results suggest that the suppression of feeding by CCK does not require an intact vasopressinergic system. The greater feeding suppression by naloxone in DI rats may suggest that opiates are interacting with vasopressin in producing their effects on food intake.     

Naloxene enhances stress-induced increases in noradrenaline turnover in specific brain regions in rats

Male Wistar rats were injected subcutaneously with either saline or naloxone, 1 mg/kg or 5 mg/kg, 10 min before exposure to 1-hour immobilization-stress. Control animals were sacrificed 70 min after respective injections. Levels of noradrenaline (NA) and its major metabolite, 3-methoxy-4-hydroxyphenylethyleneglycol sulfate (MHPG-SO₄) in seven discrete brain regions and plasma corticosterone levels were fluorometrically determined. Immobilization stress caused significant elevations of plasma corticosterone which were not affected by pretreatment with naloxone. In the hypothalamus, amygdala and thalamus, immobilization-stress caused significant elevations of MHPG-SO₄ levels, and naloxone at 5 mg/kg significantly enhanced these stress-induced elevations virtually without affecting the basal level of the metabolite. In contrast, in the hippocampus, cerebral cortex and pons plus medulla oblongata, MHPG-SO₄ levels were elevated by stress, but were not affected by naloxone pretreatment. The effect of naloxone on stress-induced reductions of NA levels was unclear, since naloxone by itself (5 mg/kg) significantly decreased the amine levels in 5 of 7 brain regions examined. These results indirectly suggest that endogenous opioid peptides in the hypothalamus, amygdala and thalamus are partly involved in the stress process and attenuate increases in NA turnover induced by stress.