α_2AAdrenergic Receptor Activation Inhibits Epileptiform Activity in the Rat Hippocampal CA3 Region

Abstract

Norepinephrine has potent antiepileptic properties, the pharmacology of which is unclear. Under conditions in which GABAergic inhibition is blocked, norepinephrine reduces hippocampal cornu ammonis 3 (CA3) epileptiform activity through ␣ 2 adrenergic receptor (AR) activation on pyramidal cells. In this study, we investigated which ␣ 2 AR subtype(s) mediates this effect. First, ␣ 2 AR genomic expression patterns of 25 rat CA3 pyramidal cells were determined using real-time single-cell reverse transcription-polymerase chain reaction, demonstrating that 12 cells expressed ␣ 2A AR transcript; 3 of the 12 cells additionally expressed mRNA for ␣ 2C AR subtype and no cells possessing ␣ 2B AR mRNA. Hippocampal CA3 epileptiform activity was then examined using field potential recordings in brain slices. The selective ␣AR agonist 6-fluoronorepinephrine caused a reduction of CA3 epileptiform activity, as measured by decreased frequency of spontaneous epileptiform bursts. In the presence of ␤AR blockade, concentration-response curves for AR agonists suggest that an ␣ 2 AR mediates this response, as the rank order of potency was 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine (UK-14304) Ն epinephrine Ͼ6-fluoronorepinephrine Ͼ norepinephrine phenylephrine. Finally, equilibrium dissociation constants (K b ) of selective ␣AR antagonists were functionally determined to confirm the specific ␣ 2 AR subtype inhibiting CA3 epileptiform activity. Apparent K b values calculated for atipamezole (1.7 nM), MK-912 (4.8 nM), BRL-44408 (15 nM), yohimbine (63 nM), ARC-239 (540 nM), prazosin (4900 nM), and terazosin (5000 nM) correlated best with affinities previously determined for the ␣ 2A AR subtype (r ϭ 0.99, slope ϭ 1.0). These results suggest that, under conditions of impaired GABAergic inhibition, activation of ␣ 2A ARs is primarily responsible for the antiepileptic actions of norepinephrine in the rat hippocampal CA3 region.The noradrenergic system is a key modulator of numerous physiological and pathological processes. Within the central nervous system (CNS), noradrenergic neurons innervate copious neural networks and regulate a number of essential neurological functions, including attention and arousal, sleep, and learning and memory (Pupo and Minneman, 2001).

Regulator of G Protein Signaling Protein Suppression of Gαo Protein-Mediated α2A Adrenergic Receptor Inhibition of Mouse Hippocampal CA3 Epileptiform Activity

“…As illustrated in Fig. 2, the rank order of potency of the three AR agonists tested in this manner revealed that (Ϫ)EPI (31 Ϯ 8.1 nM, n ϭ 45 slices) Ͼ (Ϫ)NE (150 Ϯ 45 nM, n ϭ 15 slices) ϾϾϾ (ϩ)NE (4700 Ϯ 3300 nM, n ϭ 10 slices), which is consistent with our previous results (Jurgens et al, 2007) and the order expected for ␣ARs.…”

“…We used both pharmacological and mouse genetic models to define the receptor and G protein involved in the EPI-mediated antiepileptiform activity in the hippocampus. We have confirmed the role of the ␣ 2A AR in inhibition of hippocampal CA3 epileptiform activity in mice, as shown previously by a pharmacological approach in rats (Jurgens et al, 2007). We built upon these findings by demonstrating that this involves a PTX-sensitive G i/o -type G protein.…”

“…For this particular experiment, the EC 50 value calculated from nonlinear regression analysis was 48 nM. Our previous work in rats has shown that this effect is most likely mediated by an ␣ 2 AR in the CA3 region of the hippocampus (Jurgens et al, 2007). As illustrated in Fig.…”

“…The ␣ 2A AR is the predominant ␣ 2 AR in the CNS, and it has been implicated as the primary anticonvulsant ␣ 2 AR in rat hippocampus in vitro (Jurgens et al, 2007) and in mouse in vivo (Janumpalli et al, 1998). A previous study using dopamine ␤-hydroxylase, ␣ 2A AR, and ␣ 2C AR-KO mice showed that the proconvulsant effects of ␣ 2 AR agonists were mediated by the ␣ 2A AR autoreceptor, which decreases NE release, whereas the anticonvulsant effects of ␣ 2 AR agonists were (Szot et al, 2004).…”

“…Pharmacological and molecular cloning studies have revealed the existence of three ␣ 2 AR subtypes denoted ␣ 2A , ␣ 2B , and ␣ 2C (Bylund et al, 1994). We recently showed that NE inhibits rat hippocampal CA3 epileptiform bursts through ␣ 2A AR activation (Jurgens et al, 2007). Furthermore, specific activation of ␣ 2A ARs attenuates seizures in mice elicited by chemoconvulsants (Szot et al, 2004).…”