Is 2-Hydroxypropyl-ß-cyclodextrin a Suitable Carrier for Central Administration of Δ9 -Tetrahydrocannabinol? Preclinical Evidence.

Preclinical Research Δ9 -Tetrahydrocannabinol (THC) is a hydrophobic compound that has a potent antinociceptive effect in animals after intrathecal (IT) or intracerebroventricular (ICV) administration. The lack of a suitable solvent precludes its IT administration in humans. 2-Hydroxypropyl-ß-cyclodextrin (HPßCD) increases the water solubility of hydrophobic drugs and is approved for IT administration in humans. To investigate whether HPßCD might be a suitable carrier for ICV administration of THC in rats, two formulations containing THC complexed with HPßCD (30 and 135 µg of THC per animal) and vehicle were administered to Wistar rats. The antinociceptive effect (using the tail flick test), locomotor activity, and body temperature were evaluated. ICV injection of 135 µg of THC/HPßCD complex increased tail flick latency, reduced locomotor activity, and had a dual effect on body temperature. The 30 µg THC/HPßCD formulation only produced a hyperthermic effect. All animals appeared healthy, with no difference between the groups. These results were similar to those obtained in other preclinical studies in which THC was administered centrally using solvents that are unsuitable for IT administration in humans because of their toxicity. Our findings suggest that HPßCD may be a useful carrier for IT administration of THC in humans. Drug Dev Res 78 : 411-419, 2017. © 2017 Wiley Periodicals, Inc.

Effects of chronic Delta(9)-tetrahydrocannabinol treatment on hippocampal extracellular acetylcholine concentration and alternation performance in the T-maze.

Delta(9)-Tetrahydrocannabinol (Delta(9)-THC), the psychoactive ingredient of cannabis sativa, reduces both extracellular hippocampal acetylcholine concentration and correct alternation tasks in the T-maze. The principal aim of this study was to determine whether a chronic Delta(9)-THC treatment would induce tolerance both to the reduction of extracellular hippocampal acetylcholine concentration and memory deficit produced by the drug. Our results show that a chronic Delta(9)-THC treatment (5mg/kg, i.p., twice daily for two weeks) did not produce tolerance to the inhibitory effects induced by the drug. Moreover, no strict temporal correlation between the two Delta(9)-THC effects was observed: the inhibition in extracellular acetylcholine concentration appeared only 80 min after treatment, while the reduction of correct alternation tasks in the T-maze began after 20 min. The cognitive and cholinergic effects induced by a chronic Delta(9)-THC treatment were completely blocked by the CB(1) cannabinoid receptor antagonist SR 141716A, indicating an involvement of CB(1) cannabinoid receptors in the persistent negative effects induced by the drug. These findings confirm the proposition that CB(1) cannabinoid receptors mediate the negative effects induced by Delta(9)-THC both on hippocampal extracellular acetylcholine concentration and correct alternation tasks in the T-maze, and they indicate that these effects may be differentiated. However, the major outcome of this work is the demonstration that no tolerance to the two inhibitory effects develops after a chronic Delta(9)-THC treatment.

The synthetic cannabinoid WIN55212-2 decreases the intraocular pressure in human glaucoma resistant to conventional therapies.

The search for new ocular hypotensive agents represents a frontier of current eye research because blindness due to optic neuropathy occurs insidiously in 10% of all patients affected by glaucoma. Cannabinoids have been proposed to lower intraocular pressure by either central or peripheral effects but a specific mechanism for this action has never been elucidated. We recently demonstrated the presence of the central cannabinoid receptor (CB(1)) mRNA and protein in the human ciliary body. In the present study we show that the synthetic CB(1) receptor agonist, WIN 55212--2, applied topically at doses of 25 or 50 microg (n = 8), decreases the intraocular pressure of human glaucoma resistant to conventional therapies within the first 30 min (15 +/- 0.5% and 23 +/- 0.9%, respectively). A maximal reduction of 20 +/- 0.7% and 31 +/- 0.6%, respectively, is reached in the first 60 min. These data confirm that CB(1) receptors have direct involvement in the regulation of human intraocular pressure, and suggest that, among various classes of promising antiglaucoma agents, synthetic CB(1) receptor agonists should deserve further research and clinical development.

Potential use of medicinal plants in the treatment of alcoholism.

The present paper briefly reviews the most relevant experimental data on the reducing effect of some medicinal herbs on voluntary alcohol intake in animal models of alcoholism. Pueraria lobata, Tabernanthe iboga, Panax ginseng, Salvia miltiorrhiza and Hypericum perforatum proved to be effective in decreasing alcohol consumption. Reduction of alcohol absorption from the gastrointestinal system appears to be a common feature among most of the above plants. These data suggest that medicinal plants may constitute novel and effective pharmacotherapies for alcoholism.

Ability of baclofen in reducing alcohol intake and withdrawal severity: I--Preclinical evidence.

BACKGROUND: The similarities between the pharmacological effects of the gamma-aminobutyric acid receptor agonist, baclofen, and the alcohol-substituting agent, gamma-hydroxybutyric acid, led us to investigate whether baclofen was capable of reducing (a) ethanol withdrawal syndrome in ethanol-dependent rats and (b) voluntary ethanol intake in ethanol-preferring rats. METHODS: In experiment 1, Wistar rats were rendered physically dependent on ethanol by the repeated administration of intoxicating doses of ethanol for 6 consecutive days. Baclofen was acutely administered intraperitoneally at doses of 10, 20, and 40 mg/kg. In experiment 2, baclofen (0, 2.5, 5, and 10 mg/kg, intraperitoneally) was administered once a day for 14 consecutive days to ethanol-preferring sP rats that had continuous access to ethanol (10%, v/v) and water under the two-bottle free choice regimen. RESULTS: In experiment 1, baclofen dose-dependently decreased the intensity of ethanol withdrawal signs; furthermore, 20 mg/kg of baclofen protected from audiogenic seizures in ethanol-withdrawn rats. In experiment 2, baclofen selectively and dose-dependently reduced voluntary ethanol intake; a compensatory increase in water intake left total fluid intake virtually unchanged. CONCLUSIONS: These results are in close agreement with those of a preliminary clinical study and suggest that baclofen may constitute a novel therapeutic agent for alcoholism.

Salvia miltiorrhiza extract inhibits alcohol absorption, preference, and discrimination in sP rats.

Experiment 1 of the present study investigated the ability of a standardized extract of Salvia miltiorrhiza in reducing voluntary ethanol intake in ethanol-preferring rats of the sP line. Ethanol intake occurred under the two-bottle free-choice regimen between 10% (v/v) ethanol and water in daily 4-h scheduled access periods; water was present 24 h/day. Intragastric administration of 200 mg/kg Salvia miltiorrhiza extract resulted in approximately 40% reduction in ethanol intake and preference throughout the 4-day treatment. This effect of Salvia miltiorrhiza extract was likely due to its ability of altering ethanol absorption from the gastrointestinal tract. Indeed, Experiments 2 and 3 of this study demonstrated that 200 mg/kg Salvia miltiorrhiza extract reduced blood ethanol levels (BELs) up to 60% in comparison to control rats, when ethanol was given IG, whereas it failed to modify BELs when ethanol was injected IP. The reducing effect of Salvia miltiorrhiza extract on ethanol absorption may have therefore resulted in an attenuated perception of the psychoactive effects of ethanol sought by ethanol-drinking rats. Consistently, the results of Experiment 4 of the present study demonstrated that a combination of 200 mg/kg Salvia miltiorrhiza extract IG and 1 or 2 g/kg ethanol IG resulted in a partial blockade of the discriminative stimulus effects of ethanol in sP rats trained to discriminate these doses of ethanol from water in a drug discrimination procedure. Collectively, the results are discussed as being suggestive that drugs curbing ethanol absorption from the gastrointestinal tract may constitute a novel strategy for controlling excessive alcohol consumption in human alcoholics.

Cannabinoid receptor CB1 mRNA is highly expressed in the rat ciliary body: implications for the antiglaucoma properties of marihuana.

We used RT-PCR to measure relative differences in cannabinoid receptor (CB) mRNAs in the rat eye, comparing CB1 or CB2 transcripts to that of the normalizing reference gene beta2 microglobulin (beta2m). Significantly higher levels of CB1 mRNA levels were found in the ciliary body (0.84+/-0.05% of beta2m) than in the iris, (0.34+/-0.04% of beta2m), retina (0.07+/-0.005% of beta2m) and choroid (0.06+/-0.005% of beta2m). CB2 mRNA was undetectable. This expression pattern supports a specific role for the CB1 receptor in controlling intraocular pressure, helping to explain the antiglaucoma property of cannabinoids.

Serotonin is reduced in the frontal cortex of Sardinian ethanol-preferring rats.

Ethanol-naive Sardinian alcohol-preferring (sP) and Sardinian alcohol-non-preferring (sNP) rats were tested to evaluate the levels of serotonin (5-HT) and 5-hydroxyindol-3-yl-acetic acid (5-HIAA) in the frontal cortex, hypothalamus, and nucleus accumbens, and the levels of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in the hypothalamus and nucleus accumbens. Compared with the sNP line, the sP rats had lower 5-HT and 5-HIAA concentrations in the frontal cortex, whereas no differences were found in the other brain areas tested, neither for neurotransmitters nor their metabolites. As the decreased 5-HT function is a feature shared by different alcohol-preferring strains, it could be linked to the genetic predisposition to voluntary ethanol consumption.

Effect of apomorphine on oxytocin concentrations in different brain areas and plasma of male rats.

The effect of the dopamine (DA) agonist, apomorphine, on oxytocin concentrations in the hypothalamus, hippocampus, septum and plasma was studied in male rats. Apomorphine dose dependently increased the concentration of oxytocin in the plasma and hippocampus, the minimal effective dose being 80 micrograms/kg s.c., which induced a 65% increase in plasma and a 45% increase in the hippocampus. The maximal effect (210 and 125% above controls) was induced with 240 micrograms/kg s.c. In contrast, there was a significant decrease (32%) in the oxytocin concentration in the hypothalamus, but only after the highest doses of apomorphine, while no change was found in the septum. The apomorphine effect in the hippocampus and hypothalamus was prevented by the mixed DA D-1/D-2 receptor blocker, haloperidol (0.3 mg/kg i.p.), and by the DA D-2 receptor blocker, (-)-sulpiride (20 mg/kg i.p.), but not by the DA D-1 receptor blocker, SCH 23390 (0.2 mg/kg s.c.). Similar effects were found in plasma, although SCH 23390 inhibited the apomorphine effect by 45%. Our results suggest that apomorphine stimulates oxytocinergic transmission in male rats and provide biochemical support for the hypothesis that a DA-oxytocin link exists in the central nervous system.

Hypothalamic modulation of immunoreactive oxytocin in the rat thymus.

Immunoreactive oxytocin was determined in a peptidic extract of rat thymus by means of a highly specific radioimmunoassay combined with high pressure liquid chromatography fractionation. Rat thymus was found to contain 80 +/- 7.5 pg/g wet tissue (congruent to 0.56 pg/mg protein) of oxytocin-like immunoreactivity, which behaved like synthetic oxytocin in the radioimmunoassay and in two different high pressure liquid chromatography columns. Oxytocin concentration was increased by bilateral electrolytic lesion of the paraventricular nucleus of the hypothalamus (PVN), and by high doses of corticosterone (10 mg/kg IM for 7 days) but was not modified by low doses of corticosterone (1 mg/kg IM for 7 days) or by hypophysectomy. The results suggest that rat thymus synthesizes oxytocin and that thymic oxytocin concentration is modulated by the hypothalamus.

Suppression by gamma-hydroxybutyric acid of ethanol withdrawal syndrome in rats.

The ability of gamma-hydroxybutyric acid to suppress ethanol withdrawal syndrome was tested in male rats rendered physically dependent on ethanol by several intragastric administrations of ethanol (9-15 g/kg daily for 7 days). Gamma-hydroxybutyrate (0.25, 0.50 and 1.00 g/kg i.p.), administered 8 hr after the last ethanol dose, produced a dose-dependent inhibition of withdrawal signs such as tremors and audiogenically-induced seizures; the highest dose tested suppressed all ethanol withdrawal symptoms.

Monosodium glutamate does not alter ACTH- or apomorphine-induced penile erection and yawning.

The effect of the intracerebroventricular (ICV) injection of ACTH 1-24 (1, 5 and 10 micrograms) or the subcutaneous administration of apomorphine (20 and 80 micrograms/kg SC) on spontaneous penile erection and yawning was studied in rats treated with monosodium glutamate (MSG), a treatment that depletes hypothalamic ACTH, alpha-MSH and endorphin-like peptides. Neonatal MSG treatment failed to antagonize either apomorphine- or ACTH-induced yawning in male and female rats, or to alter the number of penile erection episodes induced by the two substances in male rats. In contrast, hypophysectomy, that does not alter the concentration of hypothalamic ACTH and alpha-MSH, caused a marked prevention of apomorphine- and ACTH-induced responses, in agreement with previous studies. The results suggest that the integrity of opiomelanotropinergic neurons in the hypothalamus is not necessary for the induction of yawning and penile erection by ACTH-derived peptides, and that apomorphine and other dopamine agonists apparently do not induce penile erection and yawning by releasing an ACTH-derived peptide in brain.

Oxytocin: an extremely potent inducer of penile erection and yawning in male rats.

The intracerebroventricular (i.c.v.) injection of oxytocin, in doses ranging from 5 to 90 ng (5-90 pmol) induced penile erection and yawning in male rats. Such response was not induced by doses of the peptide higher than 100 ng, nor by equimolar doses of i.c.v. [Arg8]vasopressin, ACTH-(1-24), alpha-MSH, rat corticotropin-releasing factor (rCRF), delta sleep-inducing peptide, neurotensin or substance P. Oxytocin-induced penile erection and yawning were prevented by atropine and morphine, but not by methylatropine or the opiate antagonist naloxone. Haloperidol, a dopamine receptor antagonist, was ineffective at low doses; it partially prevented penile erection but not yawning at high doses. Since oxytocin is present not only in the neurohypophysis but also in other brain areas, our results suggest that oxytocin is implicated in the regulation of penile erection and yawning, and provide further evidence that oxytocin acts as a neuropeptide in the central nervous system.

Oxytocin-induced penile erection and yawning: site of action in the brain.

Microinjection of oxytocin into the paraventricular nucleus of the hypothalamus or into the CA1 field of the hippocampus induced a dose-dependent increase in the number of penile erection and yawning episodes in male rats. The minimal effective dose of oxytocin injected into the paraventricular nucleus was 3 ng. This dose induced the above-mentioned behaviors in 60% of the treated rats. Doses of 9 ng or higher induced the symptomatology in more than 85% of the animals. On the other hand, when the peptide was injected into the CA1 field of the hippocampus, 9 ng bilaterally were necessary to elicit penile erection and yawning in 62% of the rats. Arg8-vasopressin, which only differs from oxytocin in two amino acids, induced penile erection and yawning when injected either into the paraventricular nucleus or into the hippocampus, but was 5-10 times less potent than oxytocin. Oxytocin injection into the lateral septum, caudate nucleus, subiculum, preoptic area, ventromedial nucleus and supraoptic nucleus, was ineffective. The powerful effect of oxytocin on the induction of yawning and penile erection, suggests a physiological role of hypothalamic and hippocampal oxytocin in the regulation of such responses.

Existence of different forms of adrenocorticotropin in rat hypothalamus.

Adrenocorticotropin (ACTH)-like immunoreactivity and bioactivity were extracted from rat hypothalamus and fractionated by high pressure liquid chromatography. Analysis of the fractions either by radioimmunoassay or bioassay (corticosteroid production from rat adrenal cells) revealed several peaks of immunoreactivity and bioactivity. Only 20-25% of total ACTH-like immunoreactivity and bioactivity eluted with the same retention time as authentic ACTH 1-39. The results suggest that different forms of ACTH exist in rat hypothalamus.

Suppression by progabide of ethanol withdrawal syndrome in rats.

Progabide, a specific and clinically used GABA receptor agonist, was tested for its ability to suppress ethanol withdrawal syndrome. Male rats were rendered physically dependent on ethanol by feeding for 12 days on a liquid diet in which ethanol isocalorically replaced dextrose. Progabide (100-400 mg/kg i.p.), administered 8 h after ethanol was withdrawn, produced a dose-related inhibition of both tremors and audiogenically induced seizures. A single dose of 400 mg/kg of progabide completely suppressed all ethanol withdrawal reactions. Seizures were more sensitive to the drug than tremors. The results support the view that a decrease in GABA transmission plays a role in ethanol withdrawal symptoms and suggest that progabide may be tested as a possible treatment of ethanol withdrawal syndrome in man.