Norepinephrine release from spinal synaptosomes: auto-alpha2 -adrenergic receptor modulation.
BACKGROUND
Clonidine produces analgesia after spinal injection by activating alpha2-adrenergic receptors. Recently, clonidine has been demonstrated to increase spinal release of norepinephrine (NE) in vivo, in contrast to that anticipated by classic presynaptic autoinhibition. The purpose of the current study was to determine if clonidine could inhibit release of NE in a preparation of spinal cord tissue lacking synaptic circuits.


METHODS
Crude synaptosomes were prepared from male Sprague-Dawley rat spinal cord, loaded with [3H]NE, and stimulated by potassium chloride to release [3H]NE. Samples were incubated with clonidine in the absence or presence of various inhibitors. To study the effect of alpha2a-adrenergic receptor subtypes, some animals were pretreated with an oligodeoxynucleotide (ODN) composed of a sense or antisense sequence to a portion of this receptor.


RESULTS
Potassium chloride produced a concentration-dependent increase in [3H]NE release, and this release was inhibited by clonidine with a concentration producing 50% maximal inhibition (IC50) of 1.3 microm. The effect of clonidine was inhibited by the alpha2-adrenergic antagonists, yohimbine and idazoxan, but not by alpha1-adrenergic, muscarinic, or opioid antagonists. Intrathecal pretreatment with antisense ODN to alpha2A-adrenergic receptors reduced alpha2A-adrenergic receptor protein expression compared with sense ODN control and also reduced clonidine-induced inhibition of [3H]NE release.


CONCLUSIONS
These data demonstrate the existence of classic autoinhibitory alpha2-adrenergic receptors in the spinal cord, probably of the alpha2Asubtype. They further suggest that clonidine-induced stimulation of spinal NE release must occur from indirect actions, presumably due to activation of a spinal circuit.
Li
X
X
Department of Anesthesiology, Wake Forest University School of Medicine, Winston-Salem, NC 27157-1009, USA.
Zhao
Z
Z
Pan
H L
HL
Eisenach
J C
JC
Paqueron
X
X
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D003000:Clonidine / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D019329:Idazoxan / Q000494:pharmacology; D008297:Male; D009638:Norepinephrine / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D018341:Receptors, Adrenergic, alpha-2 / Q000187:drug effects*; D013116:Spinal Cord / Q000187:drug effects* / Q000378:metabolism; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism; D014162:Transfection / Q000187:drug effects; D015016:Yohimbine / Q000494:pharmacology
C547770:Adra2a protein, rat; D000316:Adrenergic alpha-Agonists; D000317:Adrenergic alpha-Antagonists; D018341:Receptors, Adrenergic, alpha-2; D015016:Yohimbine; D011189:Potassium Chloride; D003000:Clonidine; D009638:Norepinephrine; D019329:Idazoxan