120P Queen Elizabeth II Conference Centre London
BPS Winter Meeting 2010

 

 

Subunit-selective GABAA receptor positive allosteric modulators in the rat formalin model

Hannah Mace, Rebecca Fish, Ian Machin, Gordon McMurray, Sarah Nickolls, Catherine Sweatman, Janet Nicholson. Pfizer PGRD, Sandwich Laboratories, United Kingdom.

 

GABAA receptor point-mutated mice studies have demonstrated a role of the α2 and α3 (α2/3) subunits in preclinical pain models (Knabl et al, 2008). These studies examine the effects of subunit-selective, positive allosteric modulators, with a range of in vitro α2/3 efficacies (Table 1), in the rat formalin model.

Table 1. In vitro efficacies of GABAA positive allosteric modulators.

Compound α1 efficacy (%) α2 efficacy (%) α3 efficacy (%) α5 efficacy (%) Reference
L-838,417 0 43 43 36 Atack, 2009
TPA023 0 11 20 5 Atack et al., 2006
TPA023B 3 30 50 37 Atack et al., 2010
‘Compound 15’ 0 43 70 40 Lewis et al., 2006

Following habituation to test arenas, male Sprague-Dawley rats (200-250g, CRUK) received compound, 100mg/kg gabapentin (positive control) or methyl cellulose vehicle control p.o.. One hour post-dose, formalin (Sigma) was injected in the dorsal surface of the right hindpaw (50ul, 2.5% solution). The number of flinches in the early (0-10 minutes) and late (11-60 minutes) phases were counted via an automated testing system (UC San Diego). Compounds were deemed efficacious if a decrease in flinches compared to vehicle group was statistically significant to p<0.05 using One-Way ANOVA. For GABAA benzodiazepine receptor occupancy (RO), vehicle, test compound or bretazenil (to determine non specific binding) were dosed. Rats received 10µCi/kg [3H] Ro 15-1788, i.v. and three minutes later brain/spinal cord were removed.

The three compounds with the highest in vitro α2/3 efficacy, which also had α5 efficacy, significantly decreased the late phase response. L-838,417 was efficacious at 3 and 30mg/kg (achieving free plasma levels corresponding to 50 and 100% RO). 3mg/kg TPA023B (100% RO) decreased flinching in both phases. Interestingly, ‘Compound 15’ had a statistically significant effect in the late phase at 0.3mg/kg (65% RO), but not at 3mg/kg (100% RO). No significant effect was observed with TPA023, the compound with the lowest in vitro α2/3 efficacy and no α5 efficacy, up to 10mg/kg (100% RO) (Table 2).

Table 2. Effects of GABAA positive allosteric modulators on formalin-induced flinching. Data are means +/-sem, n = 7-8

Compound and dose Phase Flinch number p value Flinch number in gabapentin group Flinch number in vehicle group
3mg/kg
L-838,417		 Late 672 (+/- 88.3) 0.035 537 (+/- 88.3) 945 (+/- 88.3)
30mg/kg
L-838,417		 Late 591 (+/- 88.3) 0.008 537 (+/- 88.3) 945 (+/- 88.3)
3mg/kg TPA023B Early 155 (+/- 14.2) 0.046 174 (+/- 15.2) 197 (+/- 14.2)
3mg/kg TPA023B Late 538 (+/- 55.2) 0.009 549 (+/- 59.0) 755 (+/- 55.2)
0.3mg/kg ‘Compound 15’ Late 709 (+/- 91.9) 0.019 550 (+/- 85.9) 1017 (+/- 85.9)
3mg/kg ‘Compound 15’ Late 875 (+/- 85.9) 0.249 550 (+/- 85.9) 1017 (+/- 85.9)
10mg/kg TPA023 Late 790 (+/- 83.1) 0.078 553 (+/- 83.1) 1004 (+/- 83.1)

These data suggest positive allosteric modulation of the GABAA receptor via the α2, α3 and/or α5 subunits decreases formalin-evoked flinching behaviour in the rat. The model discriminated TPA023, which has no α5 efficacy and lower α2/3 efficacy, from the other compounds tested. However, no clear occupancy-response relationship was obtained in the case of the other compounds tested.

 

Atack, (2009). Advances in Pharmacol. 57, 138-185.

Atack et al., (2006). J. Pharmacol. Exp. Ther. 316 (1), 410-422.

Atack et al., (2010). J. Psychopharmacol. (epub ahead of print).

Knabl et al., (2008). Nature 451 (7176), 330-334.

Lewis et al., (2006). J. Med. Chem. 49 (8), 2600-2610.