Figure 6.

ERK inhibition recapitulates the effects of Hsp90 inhibition on anti-nociception. Tail-flick, HIV neuropathy/Von Frey, and paw-incision/Von Frey tests were performed as described in the legend for Fig. 3 and under “Experimental procedures.” Data are reported as the mean ± S.D. A, vehicle or 17-AAG (0.5 nmol) combined with pifithrin-μ (50 μg), as well as single injection controls, were injected i.c.v. with a 24-h recovery period. 0.1 nmol of DAMGO was then injected i.c.v., and tail-flick latencies were determined in a time course. ** and ***, p < 0.01 and 0.001 for vehicle versus pifithrin/17-AAG; #, ##, and ###, p < 0.05, 0.01, and 0.001, respectively, for pifithrin versus pifithrin/17-AAG group; both at the same time point using a two-way ANOVA with Fisher's least significant difference post-hoc test. AUC analysis revealed: 17-AAG, 23.7% decrease; pifithrin, 0.5% increase; pifithrin/17-AAG, 21.3% decrease. Pifithrin had no effect alone or in combination with 17-AAG on tail-flick anti-nociception. The sample size of individual mice reported consisted of two technical replicates by the same experimenter on different days. B, vehicle or 5 μg of U0126 was injected i.c.v. with a 15-min treatment period followed by 0.1 nmol of i.c.v. DAMGO and a tail-flick time course. *, p < 0.05 versus same time point via two-way ANOVA with a Fisher's least significant difference post-hoc test. AUC analysis revealed a decrease of 23.5% with U0126, very similar to the AUC values for 17-AAG treatment (Table 2). The sample size of individual mice reported consisted of two technical replicates performed on different days by the same experimenter. C, paw-incision surgery and pre- and post-surgery mechanical withdrawal thresholds were performed in mice. 5 μg of U0126 or vehicle control was injected i.c.v. with a 15-min treatment time. Morphine (1 mg/kg s.c.) was then injected, and withdrawal thresholds were determined over a time course. ** and ****, p < 0.01 and 0.0001 versus same time point via two-way ANOVA with a Fisher's Least Significant Difference post-hoc test. AUC analysis revealed that U0126 reduced anti-nociception by 89.2%, very similarly to the reduction caused by 17-AAG above (Table 2). The sample size of individual mice reported consisted of 2 technical replicates performed by the same experimenter on different days. D, HIV neuropathy was induced as in Fig. 3 and under “Experimental procedures.” On day 21, U0126 or vehicle control was i.c.v. injected as above with a 15 min treatment time. This was followed by 10 mg/kg morphine s.c., and mechanical thresholds were measured in a time course. *, **** = p < 0.05, 0.0001 versus same time point by 2 Way ANOVA with Fisher's least significant difference post-hoc test. AUC analysis revealed that U0126 reduced morphine anti-nociception by 99.7%, very similar to the value for 17-AAG (Table 2). The sample size of individual mice reported consisted of three technical replicates performed by the same experimenter on different days. Both hind paws showed very similar results; only the left paw is reported here (data not shown).