Prevention of cannabinoid withdrawal syndrome by lithium: involvement of oxytocinergic neuronal activation

Abstract

Cannabis (i.e., marijuana and cannabinoids) is the most commonly used illicit drug in developed countries, and the lifetime prevalence of marijuana dependence is the highest of all illicit drugs in the United States. To provide clues for finding effective pharmacological treatment for cannabis-dependent patients, we examined the effects and possible mechanism of lithium administration on the cannabinoid withdrawal syndrome in rats. A systemic injection of the mood stabilizer lithium, at serum levels that were clinically relevant, prevented the cannabinoid withdrawal syndrome. The effects of lithium were accompanied by expression of the cellular activation marker Fos proteins within most oxytocin-immunoreactive neurons and a significant increase in oxytocin mRNA expression in the hypothalamic paraventricular and supraoptic nuclei. Lithium also produced a significant elevation of oxytocin levels in the peripheral blood. We suggest that the effects of lithium against the cannabinoid withdrawal syndrome are mediated by oxytocinergic neuronal activation and subsequent release and action of oxytocin within the CNS. In support of our hypothesis, we found that the effects of lithium against the cannabinoid withdrawal syndrome were antagonized by systemic preapplication of an oxytocin antagonist and mimicked by systemic or intracerebroventricular injection of oxytocin. These results demonstrate that oxytocinergic neuronal activation plays a critical role in the action of lithium against the cannabinoid withdrawal syndrome in rats, thus providing a potentially novel strategy for the treatment of cannabis dependence in humans.

Figures

Fig. 1.

Summed cannabinoid withdrawal scores following different treatments. A, AM281 alone induced a mild behavioral change (▴) and severe abstinence symptoms after daily HU210 injection (Δ). Daily HU210 (○) or an acute lithium injection alone (▪) did not produce abstinence symptoms. B, In comparison with saline (■) and 1 meq/kg lithium injection (▪) before AM281 precipitation, 2 meq/kg (○) inhibited (p < 0.05) and 4 (●), 8 (Δ), and 16 (▴) meq/kg blocked (p < 0.0001) the withdrawal syndrome. Data are mean ± SEM (n = 4 per group).

Fig. 2.

Fos immunoreactivity in the hypothalamus (n = 4). Microphotographs show Fos expression in the paraventricular (A, C, E, G, H) and supraoptic (B, D, F) nuclei. There was no obvious Fos expression in the hypothalamus after lithium treatment at 1 meq/kg dosage (A, B), saline injection (G), and an acute AM281 injection (H). Injection with 4 meq/kg lithium induced Fos expression in all parts of the paraventricular (C) and supraoptic (D) nuclei. AM281-precipitated cannabinoid withdrawal evoked obviously lower density of Fos-positive cells in the ventrolateral subnuclear region in the paraventricular nucleus (E, dashed area) as well as in the dorsal part of the supraoptic nucleus (F, dashed area) in comparison with the same areas inC and D (dashed areas), although other parts of these two nuclei also contained Fos immunoreactivity. Magnifications: A, C,E, G, H, 160×;B, D, F, 320×.

Fig. 3.

Microphotographs showing double-immunofluorescent labeling revealed by a confocal microscopy. Fos (green) and oxytocin (red) immunostaining was located in the same individual neurons (yellow) in the ventromedial (A) and posterior parts (B) of the paraventricular nucleus. C, Double labeling was also present in neurons in the nucleus circularis located between the paraventricular and supraoptic nuclei. V inA indicates the third ventricle, and dashed area in C indicates a blood vessel. Magnification: 400×.

Fig. 4.

Hypothalamic oxytocin mRNA expression after different treatments (n = 4 per group).A–F, Microphotographs showing the relative density and distribution of oxytocin mRNA expression in the paraventricular nucleus after saline injection (saline), lithium treatment before AM281-precipitated cannabinoid withdrawal (HU + Li + AM), lithium alone (Li), AM281-precipitated cannabinoid withdrawal (HU + AM), twice daily HU210 (HU), and a single AM281 injection (AM). Magnification: 280×. G, Densitometry of oxytocin mRNA in the hypothalamic paraventricular (PVN) and supraoptic nuclei (SON). *Indicates a significant difference in comparison with each of the three groups labeled with * (p < 0.001) as well as with each of the other three groups without the label * (p < 0.001).

Fig. 5.

Oxytocin concentrations in the plasma after different treatments (n = 4 per group). AM281-precipitated cannabinoid withdrawal with (■) or without (Δ) lithium treatment (4 meq/kg) and an acute lithium injection alone (●) rapidly and significantly elevated the plasma oxytocin levels, in comparison with saline injection (■), twice daily injection of HU210 (○), and an acute AM281 injection (▴), which produced similarly low plasma levels of oxytocin (p > 0.05). * indicates p < 0.05; **p < 0.0001 in comparison with saline injection.

Fig. 6.

Effects of oxytocin antagonistl-368,899 on the action of lithium against the cannabinoid withdrawal syndrome. Five groups of six rats each were given different treatments (see Table 1 and Materials and Methods). *Indicates a significant difference (p < 0.05) in comparison with each of the other four groups; **indicates a significant difference (p < 0.001) in comparison with ▴ (HU210 + saline + lithium + AM210) or 79 (vehicle +l-368,899 + saline + vehicle).

Fig. 7.

Effects of oxytocin on the cannabinoid withdrawal syndrome after different treatments (n = 6 per group) (see Table 1 and Materials and Methods for treatment protocols). *Indicates a significant difference (p < 0.001) in comparison with each of the other three groups without the label *.