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 α2 adrenergic receptor (AR) activation on pyramidal cells. In this study, we investigated which α2AR subtype(s) mediates this effect. First, α2AR 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 α2AAR transcript; 3 of the 12 cells additionally expressed mRNA for α2CAR subtype and no cells possessing α2BAR mRNA. Hippocampal CA3 epileptiform activity was then examined using field potential recordings in brain slices. The selective αAR agonist 6-fluoronorepinephrine caused a reduction of CA3 epileptiform activity, as measured by decreased frequency of spontaneous epileptiform bursts. In the presence of βAR blockade, concentration-response curves for AR agonists suggest that an α2AR mediates this response, as the rank order of potency was 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine (UK-14304) ≥ epinephrine >6-fluoronorepinephrine > norepinephrine ⋙ phenylephrine. Finally, equilibrium dissociation constants (Kb) of selective αAR antagonists were functionally determined to confirm the specific α2AR subtype inhibiting CA3 epileptiform activity. Apparent Kb 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 α2AAR subtype (r = 0.99, slope = 1.0). These results suggest that, under conditions of impaired GABAergic inhibition, activation of α2AARs is primarily responsible for the antiepileptic actions of norepinephrine in the rat hippocampal CA3 region.
Footnotes
-
This investigation was supported in part by North Dakota Experimental Program to Stimulate Competitive Research (ND EPSCoR) through the National Science Foundation (NSF) grants EPS-0132289 and EPS-0447679 (to V.A.D.), NSF Faculty Early Career Development (Career) Award 0347259 (to V.A.D.), National Institutes of Health grant 2P20-RR016471 from the Biomedical Research Infrastructure Networks (BRIN) program and National Institutes of Health grant 5P20-RR017699 from the Centers of Biomedical Research Excellence (COBRE) program (to V.A.D. and J.E.P.). Additional student support was provided by an Epilepsy Foundation Predoctoral Research Training Fellowship (to C.W.J.), an American Epilepsy Society Predoctoral Research Training Fellowship (to K.L.H.), a Doctoral Dissertation Assistantship from ND EPSCoR (to K.L.H.), an Advanced Undergraduate Research Award (AURA) from ND EPSCoR (to S.J.B), an Explorations in Biomedicine Under graduate Summer Research Fellowship for Native Americans from the American Physiological Society (to K.L.D.), and an Undergraduate Summer Research Training Fellowship from the University of North Dakota Office of the Associate Vice President for Medical Research (to B.L.G. and J.A.L.).
-
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
-
doi:10.1124/mol.106.031773.
-
A preliminary report of these findings was presented at the 2005 annual meeting of the Society for Neuroscience; November 12–16, 2005; Washington DC, and the 2006 annual meeting of the American Society for Pharmacology and Experimental Therapeutics, Neuropharmacology Session, April 1–5, San Francisco, CA.
-
J.E.P. and V.A.D contributed equally to this work.
-
ABBREVIATIONS: CNS, central nervous system; CA, cornu ammonis; NE, norepinephrine; LTP, long-term potentiation; EPI, epinephrine; AR, adrenergic receptor; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid; MK-912, (2S-trans)-1,3,4,5′,6,6′,7,12b-octahydro-1′,3′-dimethyl-spiro[2H-benzofuro[2,3-a]quinolizine-2,4′(1′H)-pyrimidin]-2′(3′H)-one hydrochloride; ARC-239, 2-[2-(4-(2-methoxyphenyl)piperazin-1-yl-)ethyl]-4,4-dimethyl-1,3-(2H,4H)-isoquinolindione dihydrochloride; BRL-44408, 2-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-2,3-dihydro-1-methyl-1H-isoindole maleate; ACSF, artificial cerebrospinal fluid; RT-PCR, reverse transcription polymerase chain reaction; 6FNE, 6-fluoronorepinephrine; PHE, (R)-(–)-phenylephrine; UK-14304, 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine.
-
↵1 Current affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, North Dakota State University, Fargo, North Dakota.
- Received October 13, 2006.
- Accepted March 6, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
MolPharm articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|
