The CRF1 receptor mediates the excitatory actions of corticotropin releasing factor (CRF) in the developing rat brain: in vivo evidence using a novel, selective, non-peptide CRF receptor antagonist

Abstract

Corticotropin releasing factor (CRF) is the key coordinator of the neuroendocrine and behavioral responses to stress. In the central nervous system, CRF excites select neuronal populations, and infusion of CRF into the cerebral ventricles of infant rats produces severe age-dependent limbic seizures. These seizures, like other CRF effects, result from activation of specific receptors. Both of the characterized members of the CRF receptor family (CRF1 and CRF2), are found in the amygdala, site of origin of CRF-induced seizures, and may therefore mediate these seizures. To determine which receptor is responsible for the excitatory effects of CRF on limbic neurons, a selective, non-peptide CRF1 antagonist was tested for its ability to abolish the seizures, in comparison to non-selective inhibitory analogues of CRF. Pretreatment with the selective CRF1 blocker (NBI 27914) increased the latency and decreased the duration of CRF-induced seizures in a dose-dependent manner. The higher doses of NBI 27914 blocked the behavioral seizures and prevented epileptic discharges in concurrent electroencephalograms recorded from the amygdala. The selective CRF1 blocker was poorly effective when given systemically, consistent with limited blood-brain barrier penetration. Urocortin, a novel peptide activating both types of CRF receptors in vitro, but with preferential affinity for CRF2 receptors in vivo, produced seizures with a lower potency than CRF. These limbic seizures, indistinguishable from those induced by CRF, were abolished by pretreatment with NBI 27914, consistent with their dependence on CRF1 activation. In summary, CRF induces limbic seizures in the immature rat, which are abolished by selective blocking of the CRF1 receptor. CRF1-messenger RNA levels are maximal in sites of seizure origin and propagation during the age when CRF is most potent as a convulsant. Taken together, these facts strongly support the role of the developmentally regulated CRF1 receptor in mediating the convulsant effects of CRF in the developing brain.

Fig. 1

Dose effect of pretreatment with NBI 27914 on the duration of seizures evoked by CRF. Rat pups received the selective CRF₁ inhibitor at the doses indicated on the x-axis (see Section 2.3.1). Values are means ± S.E.M. *Significantly different from control values, P-0.05; ** P-0.01 (Mann–Whitney U-test).

Fig. 2

Effects of pretreatment with NBI 27914 on EEG features of seizures induced by CRF. Six immature rats were implanted with bipolar electrodes directed at the left amygdala on the 10th postnatal day (see text). Depth EEGs were recorded on the following morning. Panel A shows low-voltage, non-rhythmic background activity in all six tracings (1–6). Occasional brief movement artifacts are noted by the arrows. Panel B was obtained 30 min after the administration of 1 mg/kg of NBI 27914 to rats Nos. 2, 3 and 6, and 13–15 min after the infusion of 0.15 nmol CRF to all six pups. The EEG features of animals Nos. 2, 3 and 6 reveal only increased overall frequency of movement artifacts. High-voltage spike-wave and polyspike epileptic discharges are evident in rats subjected to CRF without pretreatment with NBI 27914 (curved arrows). Behavioral limbic seizures lasted for 150, 180 and 110 min in rats Nos. 1, 4 and 5, respectively. No behavioral seizures were observed in rats Nos. 3 and 6. Rat No. 2 had one episode of wet dog shakes, and a seizure lasting 25 min. Horizontal marker indicates 1 s; vertical marker corresponds to 50 μV.