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Rat pineal α 1 ‐adrenoceptor subtypes: studies using radioligand binding and reverse transcription‐polymerase chain reaction analysis
Author(s) -
Sugden David,
Anwar Naveed,
Klein David C.
Publication year - 1996
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1996.tb15530.x
Subject(s) - pinealocyte , radioligand assay , radioligand , pineal gland , chemistry , binding site , oxymetazoline , endocrinology , phentolamine , medicine , microbiology and biotechnology , antagonist , receptor , biology , adrenergic receptor , biochemistry , melatonin
1 The pharmacological characteristics of α 1 ‐adrenoceptor binding sites in rat pineal gland membranes, detected by use of a selective α 1 ‐adrenoceptor antagonist ([ 125 I]‐iodo‐2‐[β‐(4‐hydroxyphenyl) ethylaminomethyl]tetralone, [ 125 I]‐HEAT), were investigated with the alkylating agent, chloroethylclonidine (CEC), and in competition experiments with a number of adrenoceptor agonists and antagonists. 2 Chloroethylclonidine (CEC) treatment (10 μ m , 10 min) of rat pineal membranes inactivated ∼ 70% of specific [ 125 I]‐HEAT binding sites. Higher concentrations of CEC (up to 100 μ m ) or longer treatment periods (upto 40 min) were no more effective. 3 Adrenoceptor agonists and antagonists competitively inhibited [ 125 I]‐HEAT binding with Hill coefficients close to unity indicating a single α 1 ‐adrenoceptor subtype is present. The affinity ( K i of subtype selective agonists (oxymetazoline, SDZ NVI‐085) and antagonists (5‐methylurapidil, WB4101, benoxathian, phentolamine) was consistent with binding to an α 1B ‐adrenoceptor subtype. 4 The (−)− and (+)‐enantiomers of niguldipine had an equal and low affinity for α 1 ‐adrenoceptor binding sites both in untreated (log K i − 6.66 and −6.90 respectively) and CEC‐treated membranes in which ∼ 70% of sites had been inactivated (log K i − 6.41 and −6.86 respectively). This indicates that the small proportion of α 1 ‐adrenoceptors insensitive to CEC are not α 1A ‐adrenoceptors. 5 mRNA was isolated from rat pinealocytes, cDNA was synthesized and then amplified by the polymerase chain reaction with α 1 ‐adrenoceptor subtype specific primers. These experiments identified both α 1A ‐ and α 1B ‐adrenoceptor mRNA, but not α 1D ‐mRNA in rat pinealocytes, although all three adrenoceptor subtypes were readily identified in rat brain cortex. 6 These data indicate that although both α 1A ‐ and α 1B ‐adrenoceptor mRNAs are present in the pineal the major subtype of α 1 ‐adrenoceptor expressed is the α 1B .