Selective blockade and recovery of cell surface alpha 2-adrenergic receptors in human erythroleukemia (HEL) cells. Studies with the irreversible antagonist benextramine

Journal Article · Thu Dec 31 23:00:00 EST 1987 · Mol. Pharmacol.; (United States)

OSTI ID:5471652

McKernan, R M; Strickland, W R; Insel, P A

alpha 2-Adrenergic receptors are present on human erythroleukemia (HEL) cells, both on the cell surface and in a sequestered compartment. In the current study we show that benextramine, a hydrophilic irreversible antagonist, can be used to investigate alpha 2-adrenergic receptor compartmentation in these cells. In membranes prepared from HEL cells, benextramine competed for all alpha 2-adrenergic receptors ( (/sup 3/H)yohimbine sites). In intact cells, at 4 degrees, benextramine exhibited a biphasic competition curve for alpha 2-adrenergic receptors, with EC50 values of approximately 10 microM and greater than 1 mM for the high and low affinity components, respectively. We propose that the alpha 2-adrenergic receptors preferentially blocked by benextramine are those on the surface of the cell, whereas those with low affinity are sequestered receptors because: 1) only epinephrine-accessible sites are removed by prior treatment of cells with benextramine, 2) a preparation enriched with surface membranes is also enriched in receptors with a high affinity for benextramine; and 3) after blockade of cell surface receptors (54 +/- 6% of total sites, n = 7) by benextramine, the ability of the alpha 2-adrenergic agonists epinephrine and UK-14,304 to inhibit forskolin-stimulated cAMP accumulation is lost. The latter result implies that only cell surface and not sequestered receptors are functionally coupled to adenylate cyclase. The return of receptors from the sequestered compartment to the cell surface and the recovery of alpha 2-adrenergic receptor function were measured after HEL cells were treated with benextramine (50 microM for 1 hr at 4 degrees). The recovery of receptor binding (t1/2 = 25 min) was somewhat slower than the recovery of function (t1/2 approximately 8 min).

• Research Organization: Univ. of California, La Jolla
• OSTI ID: 5471652
• Journal Information: Mol. Pharmacol.; (United States), Journal Name: Mol. Pharmacol.; (United States) Vol. 33:1; ISSN MOPMA
• Country of Publication: United States
• Language: English

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550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINES
AMP
ANIMAL CELLS
ANIMALS
AUTONOMIC NERVOUS SYSTEM AGENTS
BIOCHEMICAL REACTION KINETICS
CELL CONSTITUENTS
CELL MEMBRANES
CYCLASES
CYSTAMINE
DRUGS
ENZYMES
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
LYASES
MAMMALS
MAN
MEMBRANE PROTEINS
MEMBRANES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PRIMATES
PROTEINS
RADIOPROTECTIVE SUBSTANCES
REACTION KINETICS
RECEPTORS
SYMPATHOLYTICS
SYMPATHOMIMETICS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
TUMOR CELLS
VERTEBRATES