Transiently increased glutamate cycling in rat PFC is associated with rapid onset of antidepressant-like effects

Abstract

Several drugs have recently been reported to induce rapid antidepressant effects in clinical trials and rodent models. Although the cellular mechanisms involved remain unclear, reports suggest that increased glutamate transmission contributes to these effects. Here, we demonstrate that the antidepressant-like efficacy of three unique drugs, with reported rapid onset antidepressant properties, is coupled with a rapid transient rise in glutamate cycling in the medial prefronal cortex (mPFC) of awake rats as measured by ex vivo ¹H-[¹³C]-nuclear magnetic resonance spectroscopy. Rats were acutely pretreated by intraperitoneal injection with a single dose of ketamine (1, 3, 10, 30 and 80 mg kg^-1), Ro 25-6981 (1, 3 and 10 mg kg^-1), scopolamine (5, 25 and 100 μg kg^-1) or vehicle (controls). At fixed times after drug injection, animals received an intravenous infusion of [1,6-¹³C₂]glucose for 8 min to enrich the amino-acid pools of the brain with ¹³C, followed by rapid euthanasia. The mPFC was dissected, extracted with ethanol and metabolite ¹³C enrichments were measured. We found a clear dose-dependent effect of ketamine and Ro 25-6981 on behavior and the percentage of ¹³C enrichment of glutamate, glutamine and GABA (γ-aminobutyric acid). Further, we also found an effect of scopolamine on both cycling and behavior. These studies demonstrate that three pharmacologically distinct classes of drugs, clinically related through their reported rapid antidepressant actions, share the common ability to rapidly stimulate glutamate cycling at doses pertinent for their antidepressant-like efficacy. We conclude that increased cycling precedes the antidepressant action at behaviorally effective doses and suggest that the rapid change in cycling could be used to predict efficacy of novel agents or identify doses with antidepressant activity.

Figure 1. Effects of ketamine and Ro 25-6981 on mPFC glutamate/GABA-glutamine metabolism and FST behavior

(A) Percent change in PFC glutamate-C4, GABA-C2, and glutamine-C4 fractional enrichments over an interval from 10 to 18min after i.p. doses of 1, 3, 10, 30 and 80mg/kg compared to vehicle-injected animals. (B) Ketamine 30mg/kg decreased average time immobile in the forced swim test (FST) but not 3 and 10mg/kg. The effect was more pronounced in the last 5min of the test. (C) Percent change in PFC glutamate-C4, GABA-C2, and glutamine-C4 enrichments over an interval from 30 to 38min after i.p. doses of 1, 3 and 10mg/kg Ro 25-6981 relative to vehicle-treated controls. (D) Averaged over the two blocks of 5min, the Ro 25-6981 10mg/kg animal group shows a trend for decreased time immobile with a more pronounced effect for the last 5min (#p_unadj=0.05 p_Bonferroni =0.1). Interrogation time is the 8min interval after ketamine injection during which the ¹³C-glucose was infused. (#p<0.1 *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001).

Figure 2. Effects of time after ketamine injection on glutamate, GABA and glutamine ¹³C labeling compared to control animals

(A) The figure depicts the percent change in fractional enrichment compared to vehicle-injected animals over time. Amino acid labeling reflects the interval from 0.5 to 8.5min, 10 to 18min, 30 to 38min, and 60 to 68min after i.p. injection of ketamine 30mg/kg. (B) There was no significant effect of a single 10mg/kg i.p. dose of ketamine on glutamate, GABA and glutamine ¹³C labeling from ¹³C-glucose when interrogated for 8mins at 24h. (#p<0.1; *p<0.05; **p<0.01; ***p<0.001).

Figure 3. Effects of Scopolamine on FST immobility and glutamate, GABA and glutamine ¹³C labeling compared to control animals

(A) Scopolamine decreased average time immobile in the FST at 25μg/kg but not 5 and 100μg/kg. Averaged over the two blocks of 5 min, there was a significant interaction between drug and time (F3,48=4.03, p<0.05) with a more pronounced effect with scopolamine (Scop) 25μg/kg for the last 5 min (*p_Bonferroni <0.05) when compared to the vehicle-injected animal group. (B) Effects of a single 25μg/kg i.p. dose of scopolamine on magnitude change of glutamate, GABA and glutamine ¹³C labeling from glucose over control (*p <0.05 when compared to vehicle-injected animal group).