Characterization of the Visceral Antinociceptive Effect of Glial Glutamate Transporter GLT-1 Upregulation by Ceftriaxone

Abstract

Recent studies demonstrate that glial glutamate transporter-1 (GLT-1) upregulation attenuates visceral nociception. The present work further characterized the effect of ceftriaxone- (CTX-) mediated GLT-1 upregulation on visceral hyperalgesia. Intrathecal pretreatment with dihydrokainate, a selective GLT-1 antagonist, produced a reversal of the antinociceptive response to bladder distension produced by CTX. The hyperalgesic response to urinary bladder distension caused by intravesicular acrolein was also attenuated by CTX treatment as was the enhanced time spent licking of abdominal area due to intravesicular acrolein. Bladder inflammation via cyclophosphamide injections enhanced the nociceptive to bladder distension; cohorts administered CTX and concomitant cyclophosphamide showed reduced hyperalgesic response. Cyclophosphamide-induced bladder hyperalgesia correlated with a significant 22% increase in GluR1 AMPA receptor subunit expression in the membrane fraction of the lumbosacral spinal cord, which was attenuated by CTX coadministration. Finally, neonatal colon insult-induced hyperalgesia caused by intracolonic mustard oil (2%) administration at P9 and P11 was attenuated by CTX. These studies suggest that GLT-1 upregulation (1) attenuates the hyperalgesia caused by bladder irritation/inflammation or by neonatal colonic insult, (2) acts at a spinal site, and (3) may produce antinociceptive effects by attenuating GluR1 membrane trafficking. These findings support further consideration of this FDA-approved drug to treat chronic pelvic pain syndromes.

Figure 1

(a)–(c) Intrathecal injection of dihydrokainate (DHK), a selective GLT-1 antagonist, reversed the attenuated visceromotor response induced by 1 wk ceftriaxone (CTX) at 0.3 mM and 0.03 mM, but not the 0.003 mM dose. (a) Mice that received 1 wk VEH + i.t. DHK (0.3 mM) showed an elevated visceromotor response relative to control (1 wk VEH + i.t. VEH). Treatment with i.t. DHK (0.3 mM) reversed the CTX-attenuated visceromotor response. (b) Mice treated with 1 wk VEH + i.t. DHK (0.03 mM) showed no difference in the visceromotor response relative to control (P > 0.05). Treatment with i.t. DHK (0.03 mM) reversed the CTX-attenuated visceromotor response. (c) In contrast, mice administered 1 wk CTX + i.t. DHK (0.003 mM) showed no reversal of the attenuated visceromotor response produced by 1 wk CTX + i.t. VEH.

Figure 2

Lower doses of CTX (50 mg/kg or 100 mg/kg × 7 days) did not produce a visceromotor response different from the control group at graded distension volumes. Animals administered 1 wk VEH + intravesicular acrolein (ivc ACRO) showed a significant increase in the visceromotor response compared to the control group at 0.15 and 0.2 mL (∗P < 0.05). In contrast, mice treated with 1 wk CTX (50 or 100 mg/kg) + ivc ACRO did not show an enhanced visceromotor response to bladder distension compared to the 1 wk VEH + ivc VEH treated cohort at 0.15 and 0.2 mL.

Figure 3

Mice treated with vehicle or CTX (100 mg/kg) for seven days were observed for the amount of time spent licking the abdominal area for 1 hr after receiving intravesicular (ivc) vehicle or acrolein (0.4 mM). The data show a significantly enhanced time spent licking in animals that received 1 wk VEH + ivc ACRO, compared to the control group (1 wk VEH + ivc VEH; ∗P < 0.05). This enhanced nociceptive response was attenuated in cohorts receiving 1 wk CTX + ivc ACRO treatment.

Figure 4

(a)-(b) Cyclophosphamide (1 wk VEH + 1 wk CYP) significantly increased the visceromotor to urinary bladder distension at 0.15 and 0.2 mL, compared to 1 wk VEH + 1 wk VEH controls. (∗P < 0.05). (a) The enhanced visceromotor response produced in the 1 wk VEH + CYP group was attenuated by 1 wk daily CTX administration (1 wk CTX (200 mg/kg) + 1 wk CYP). (b) Administration of a lower dose of CTX (100 mg/kg; i.p.) for seven days also attenuated the enhanced visceromotor response to bladder distension of mice treated with cyclophosphamide (80 mg/kg) during the 7-day period prior to undergoing bladder distension (∗P < 0.05).

Figure 5

(a)-(b) Total and membrane-associated GluR1 is increased after 1-week cyclophosphamide treatment. Enhanced GluR1 membrane trafficking is mitigated after concomitant 1 wk CTX and cyclophosphamide treatment. (a) Western blot comparing Total, cytosolic, and membrane GruR1 expression. (b) Representative western blot densitometry shows a 20% increase in total GluR1 expression in 1 wk VEH + CYP and a 22% increase in the membrane compared to control cohorts 1 wk VEH + VEH (∗P < 0.05).

Figure 6

Neonatal stress-induced visceral hypersensitivity in adult mice. Postnatal day 9 and 11 treatment with intracolonic 2% mustard oil (1 wk VEH + i.c. MO) dramatically increased the visceromotor response to colorectal distension at 30–60 mm Hg in adult mice, compared to control (1 wk VEH + i.c. VEH) cohorts. In marked contrast, ceftriaxone administered 1-week before colon distension (1 wk CTX + (i.c.) MO) completely attenuated the enhanced visceromotor response to colorectal distension produced by neonatal colon irritation (∗P < 0.05).