Betaxolol, a selective beta(1)-adrenergic receptor antagonist, diminishes anxiety-like behavior during early withdrawal from chronic cocaine administration in rats

Abstract

Background: Anxiety has been indicated as one of the main symptoms of the cocaine withdrawal syndrome in human addicts and severe anxiety during withdrawal may potentially contribute to relapse. As alterations in noradrenergic transmission in limbic areas underlie withdrawal symptomatology for many drugs of abuse, the present study sought to determine the effect of cocaine withdrawal on beta-adrenergic receptor (beta(1) and beta(2)) expression in the amygdala.

Methods: Male Sprague Dawley rats were administered intraperitoneal (i.p.) injections of cocaine (20 mg/kg) once daily for 14 days. Two days following the last cocaine injection, amygdala brain regions were micro-dissected and processed for Western blot analysis. Results showed that beta(1)-adrenergic receptor, but not beta(2)-adrenergic receptor expression was significantly increased in amygdala extracts of cocaine-withdrawn animals as compared to controls. This finding motivated further studies aimed at determining whether treatment with betaxolol, a highly selective beta(1)-adrenergic receptor antagonist, could ameliorate cocaine withdrawal-induced anxiety. In these studies, betaxolol (5 mg/kg via i.p. injection) was administered at 24 and then 44 h following the final chronic cocaine administration. Anxiety-like behavior was evaluated using the elevated plus maze test approximately 2 h following the last betaxolol injection. Following behavioral testing, betaxolol effects on beta(1)-adrenergic receptor protein expression were examined by Western blotting in amygdala extracts from rats undergoing cocaine withdrawal.

Results: Animals treated with betaxolol during cocaine withdrawal exhibited a significant attenuation of anxiety-like behavior characterized by increased time spent in the open arms and increased entries into the open arms compared to animals treated with only saline during cocaine withdrawal. In contrast, betaxolol did not produce anxiolytic-like effects in control animals treated chronically with saline. Furthermore, treatment with betaxolol during early cocaine withdrawal significantly decreased beta(1)-adrenergic receptor protein expression in the amygdala to levels comparable to those of control animals.

Conclusions: The present findings suggest that the anxiolytic-like effect of betaxolol on cocaine-induced anxiety may be related to its effect on amygdalar beta(1)-adrenergic receptors that are up-regulated during early phases of drug withdrawal. These data support the efficacy of betaxolol as a potential effective pharmacotherapy in treating cocaine withdrawal-induced anxiety during early phases of abstinence.

Figure 1. β₁–adrenergic receptor expression is increased following early (2 day) withdrawal from chronic cocaine administration

Amygdala extracts from rats that were euthanized following chronic cocaine administration (20 mg/kg i.p. for 14 days) with no withdrawal period exhibited no significant difference in the expression of either β₁ or β₂ -adrenergic receptors compared to saline-treated animals. Following 2 day withdrawal from chronic cocaine administration, β₁–adrenergic receptor expression was significantly increased (p < 0.05), while β₂-adrenergic receptor expression was unchanged in rat amygdala extracts. Animals that underwent 12 days of withdrawal following chronic cocaine administration exhibited no significant change in amygdala protein expression levels of either β₁ or β₂ -adrenergic receptors.

Figure 2. Dose response of betaxolol administration on cocaine withdrawal-induced anxiety-like behavior measured by the elevated plus maze

Data is represented as (A) the mean percentage of time spent in the open arms/total time spent in both open + closed arms (± S.E.M) and (B) the mean percentage of the entries into the open arms/total number of entries into both the open and closed arms (± S.E.M.). Prior to testing, rats were administered chronic cocaine (20 mg/kg i.p. daily for 14 days) followed by one of three different betaxolol treatments (0.5, 2.0 or 5.0 mg/kg i.p.) at 24 and 44 hours following the last cocaine injection. Control animals received either chronic i.p. injections of cocaine (20 mg/kg) or saline (1 ml/kg) for 14 days followed by administration of saline (1 ml/kg) at 24 and 44 hours during the withdrawal phase. (*p < 0.05)

Figure 3. Betaxolol diminishes anxiety-like behavior during early abstinence from chronic cocaine administration in rats

Data is represented as (A) the mean percentage of time spent in the open arms/total time spent in both open + closed arms (± S.E.M); (B) the mean percentage of the entries into the open arms/total number of entries into both the open and closed arms (± S.E.M.); and (C) the mean number of closed arm entries (± S.E.M). Prior to testing, rats were administered either chronic i.p. injections of cocaine (20 mg/kg) or saline (1 ml/kg) for 14 days followed by either i.p. betaxolol (5 mg/kg) or saline (1 ml/kg) at 24 and 44 hours following the last drug injection. (*p < 0.05, **p < 0.01, ***p < 0.001)

Figure 4. Anxiolytic-like effects of betaxolol demonstrated during early withdrawal from chronic cocaine administration are not attributable to alterations in ambulatory activity

Data is represented as (A) the mean distance traveled per minute of the testing session (± S.E.M) and (B) the mean average velocity (in centimeters per second) per minute of the testing session (± S.E.M). For each animal, the mean of the distance traveled was calculated from the total distance traveled over the ten minute testing session that was reported in one minute increments. The mean of the average velocity for each animal was calculated from the summation of the average velocities recorded in one minute increments over the ten minute testing session..

Figure 5. Betaxolol treatment during early cocaine withdrawal decreases β₁–adrenergic receptor protein expression in the amygdala to levels comparable to that of control animals

β₁-adrenergic receptor immunoreactivity was evaluated in amygdala and frontal cortex extracts from animals that were administered either chronic i.p. injections of cocaine (20 mg/kg) or saline (1 ml/kg) for 14 days followed by either i.p. betaxolol (5 mg/kg) or saline (1 ml/kg) at 24 and 44 hours following the last drug injection. β₁-adrenergic receptor immunoreactivity in the amygdala of experimental animals is expressed as a percentage of the control mean when the control equals 100. Significant differences are indicated: *, significantly different from saline control animals (p < 0.05) and #, significantly different than cocaine control animals.