Corticotropin-releasing factor 1 and 2 receptors in the dorsal raphé differentially affect serotonin release in the nucleus accumbens

Abstract

Corticotropin-releasing factor (CRF) is a neurohormone that mediates stress, anxiety, and affects serotonergic activity. Studies have shown that CRF has dose-dependent opposing effects on serotonergic activity. This effect has been hypothesized to be differentially mediated by CRF(1) and CRF(2) receptors in the dorsal raphé nucleus. We directly tested this hypothesis by using in vivo microdialysis to determine the effects of CRF and CRF receptor antagonists in the dorsal raphé nucleus on serotonin (5-HT) release in the nucleus accumbens, a brain region implicated in the neuropathology of stress-related psychiatric disorders. Male urethane-anesthetized rats were implanted with a microdialysis probe into the nucleus accumbens, and CRF (0, 100 or 500 ng) was infused into the dorsal raphé. Infusion of CRF into the dorsal raphé nucleus had dose-dependent opposite effects, with 100 ng of CRF significantly decreasing 5-HT levels in the nucleus accumbens and 500 ng CRF significantly increasing accumbal 5-HT levels. In subsequent experiments, the raphé was pre-treated with the CRF(1) receptor antagonist antalarmin (0.25 microg) or the CRF(2) receptor antagonist antisauvagine-30 (ASV-30; 2 microg) prior to CRF infusion. Antagonism of CRF(1) receptors in the dorsal raphé nucleus abolished the decrease in accumbal 5-HT levels elicited by 100 ng CRF, and CRF(2) receptor antagonism in the raphé blocked the increase in accumbal 5-HT levels elicited by 500 ng CRF. These results suggest that the opposing effects of dorsal raphé CRF on 5-HT release in the nucleus accumbens are dependent on differential activation of CRF(1) and CRF(2) receptors in the dorsal raphé nucleus.

Figure 1. Probe and Cannula Placements in the Nucleus Accumbens and dorsal Raphé

Schematic representations of microdialysis probe (A,C,E) and dorsal raphé nucleus drug infusion cannulae (B, D, F) for CRF dose-response experiments (A and B), CRF₁ receptor antagonist experiments (C and D), and CRF₂ receptor antagonist experiments (E and F). Microdialysis probes (black bars which represent more than one placement) were placed in the nucleus accumbens (NAc) with a 2 mm membrane to record from both the shell and the core (A, C, E; Figures adapted from Paxinos and Watson (1997); AP: +1.2 from bregma; ML: −1.4 from midline; DV: 8.1 from dura). Drug cannulae (black circles) were placed in regions encompassing areas of the dorsal raphé nucleus (AP: −7.4 from bregma; ML: +2.8 from midline; DV: −4.6 from dura) that project to the nucleus accumbens (B, D, and F). Figure B also illustrates cannula placements outside the dorsal raphé nucleus (crosses).

Figure 2. Effects of CRF Infusion into the dorsal Raphé nucleus on Accumbal 5-HT levels

Infusion of 100 ng CRF into the dorsal raphé nucleus (A) resulted in a significant decrease in accumbal 5-HT when compared to CSF (A; *significant difference in accumbal 5-HT levels as compared to aCSF infusion, P<0.05), and compared to pre-infusion baseline 5-HT overflow (# significant difference over time in accumbal 5-HT levels, P<0.05). Arrow indicates time of intra-dorsal raphé injection. In contrast, infusion of 500 ng into the dorsal raphé nucleus (A) resulted in a significant increase in accumbal 5-HT levels when compared to controls (A). When cannulae placements were outside the dorsal raphé nucleus (B), no change in 5-HT overflow was recorded, regardless of the treatment.

Figure 3. Infusion of a CRF₁ Antagonist into the dorsal Raphé nucleus Blocks the Effects of 100 ng CRF Infusion into the dorsal Raphé

Infusion of 100 ng CRF into the dorsal raphé nucleus (in the absence of the CRF₁ antagonist antalarmin) resulted in a significant 5-HT decrease in the nucleus accumbens (* significant compared to control groups; # significant difference over time; P<0.05). The effect of 100 ng CRF on accumbal 5-HT levels was not affected by infusion of the CRF₂ antagonist (ASV-30) into the dorsal raphé nucleus 10 min prior to the infusion of 100 ng CRF. In contrast, infusion of antalarmin, a CRF₁ receptor antagonist, into the dorsal raphé nucleus completely blocked the 100 ng CRF effect. No effects on accumbal 5-HT levels were observed after infusion of vehicle or antalarmin alone in the absence of CRF. Arrows indicate time of intra-dorsal raphé injection: first arrow represents infusion of receptor antagonist or vehicle, and the second arrow represents infusion of CRF.

Figure 4. Infusion of a CRF₂ Antagonist into the dorsal Raphé nucleus Blocks the Effects of 500 ng CRF Infused into the dorsal Raphé

Infusion of 500 ng CRF into the dorsal raphé nucleus (in the absence of the CRF₂ antagonist ASV-30) resulted in a significant increase in accumbal 5-HT (* significant compared to control groups; # significant difference over time; P<0.05). The effect of 500 ng CRF on accumbal 5-HT levels was not affected by infusion of the CRF₁ antagonist (antalarmin) into the dorsal raphé nucleus 10 min prior to the infusion of 500 ng CRF. In contrast, infusion of ASV-30, blocked the effects of 500 ng CRF into the dorsal raphé nucleus. No effects on accumbal 5-HT levels were observed after infusion of vehicle or ASV-30 alone in the absence of CRF. Arrows indicate time of intra-dorsal raphé injection: first arrow represents infusion of receptor antagonist or vehicle, and the second arrow represents infusion of CRF.