Pharmacological and anatomical evidence for an interaction between mGluR5- and GABA(A) alpha1-containing receptors in the discriminative stimulus effects of ethanol

Abstract

The discriminative stimulus properties of ethanol are mediated in part by positive modulation of GABA(A) receptors. Recent evidence indicates that metabotropic glutamate receptor subtype 5 (mGluR5) activity can influence GABA(A) receptor function. Therefore, the purpose of this work was to examine the potential involvement of mGluR5 in the discriminative stimulus effects of ethanol. In rats trained to discriminate ethanol (1 g/kg, intragastric gavage (i.g.)) from water, 2-methyl-6-(phenylethyl)-pyridine (MPEP) (1-50 mg/kg, i.p.) a selective noncompetitive antagonist of the mGlu5 receptor did not produce ethanol-like stimulus properties. However, pretreatment with MPEP (30 mg/kg) reduced the stimulus properties of ethanol as indicated by significant reductions in ethanol-appropriate responding, specifically at 0.5 and 1 g/kg ethanol, and a failure of ethanol test doses (1 and 2 g/kg) to fully substitute for the ethanol training dose. To test whether mGluR5 antagonism altered the GABA(A) receptor component of the ethanol stimulus, the ability of MPEP to modulate pentobarbital and diazepam substitution for ethanol was assessed. Pentobarbital substitution (1-10 mg/kg, i.p.) for ethanol was not altered by MPEP pretreatment. However, MPEP pretreatment inhibited the ethanol-like stimulus properties of diazepam (5 mg/kg, i.p.). To examine a potential anatomical basis for these pharmacological findings, expression patterns of mGluR5- and benzodiazepine-sensitive GABA(A) alpha1-containing receptors were examined by dual-label fluorescent immunohistochemistry with visualization by confocal microscopy. Results indicated that mGluR5- and GABA(A) alpha1-containing receptors were both coexpressed in limbic brain regions and colocalized on the same cells in specific brain regions including the amygdala, hippocampus, globus pallidus, and ventral pallidum. Together, these findings suggest an interaction between mGluR5- and benzodiazepine-sensitive GABA(A) receptors in mediating ethanol discrimination.

Figure 1

(a) Mean (±SEM) percentage of ethanol-appropriate responding upon completion of the first FR10 at each ethanol dose tested (n = 12). (b) Mean (±SEM) test session response rate at each ethanol dose tested. Data points to the left of the x-axis break represent performance on the water (W) and ethanol (E) training session prior to testing. Data points to the right of the x-axis break represent test session performance following IG ethanol administration. The horizontal dashed line (>80%) represents full substitution for the discriminative stimulus effects of ethanol (1 g/kg, i.g.). *Significant difference from 0.1 g/kg ethanol (Tukey’s, p<0.05).

Figure 2

(a) Mean (±SEM) percentage of ethanol-appropriate responding upon completion of the first FR10 at each MPEP dose tested (n = 6). (b) Mean (±SEM) test session response rate at each MPEP dose tested. Data points to the left of the x-axis break represent performance on the water (W) and ethanol (E) training session prior to testing. Data points to the right of the x-axis break represent test session performance following i.p. MPEP administration. The horizontal dashed line (>80%) represents full substitution for the discriminative stimulus effects of ethanol (1 g/kg, i.g.). *Significant difference from saline (Tukey’s, p<0.05).

Figure 3

(a) Mean (±SEM) percentage of ethanol-appropriate responding upon completion of the first FR10 after saline or MPEP pretreatment at each ethanol dose (n = 8). (b) Mean (±SEM) test session response rate after saline or MPEP pretreatment at each ethanol dose tested. Data points to the left of the x-axis break represent performance on the water (W) and ethanol (E) training session prior to testing. The horizontal dashed line (>80%) represents full substitution for the discriminative stimulus effects of ethanol (1 g/kg, i.g.). *Significantly different from the respective 0.1 g/kg ethanol dose within pretreatment condition. ^†Significant difference between saline and MPEP pretreatment (Tukey’s, p>0.05).

Figure 4

(a) Mean (±SEM) percentage of ethanol-appropriate responding upon completion of the first FR10 after saline or MPEP pretreatment at each pentobarbital dose tested (n = 10). (b) Mean (±SEM) test session response rate after saline or MPEP pretreatment at each pentobarbital dose tested. Data points to the left of the x-axis break represent performance on the water (W) and ethanol (E) training session prior to testing. The horizontal dashed line (>80%) represents full substitution for the discriminative stimulus effects of ethanol (1 g/kg, i.g.). *Significantly different from the respective 1 mg/kg pentobarbital dose within pretreatment condition. ^†Significant difference between saline and MPEP pretreatment (Tukey’s, p<0.05).

Figure 5

(a) Mean (±SEM) percentage of ethanol-appropriate responding upon completion of the first FR10 after saline or MPEP pretreatment at each diazepam dose tested (n = 11). (b) Mean (±SEM) test session response rate after saline or MPEP pretreatment at each diazepam dose tested. Data points to the left of the x-axis break represent performance on the water (W) and ethanol (E) training session prior to testing. The horizontal dashed line (>80%) represents full substitution for the discriminative stimulus effects of ethanol (1 g/kg, i.g.). *Significantly different from the respective 1 mg/kg diazepam dose within pretreatment condition. ^†Significant difference between saline and MPEP pretreatment (Tukey’s, p<0.05).

Figure 6

Confocal images from a representative rat showing the α1-subunit of the GABA_A receptor and mGluR5 in the core of the nucleus accumbens (AcbC; a–c), the CA3 region of the hippocampus (CA3; d–f), the ventral pallidum (VP; g–i), and the globus pallidus (GP; j–l). GABA_A α1 labeling appears in red, and mGluR5 labeling appears in green. Images a–i taken at × 63 magnification; images j–l taken at × 40 magnification.