Abstract
A new adenosine analogue, (-)-iodo-N6-phydroxyphenylisopropyladenosine
(-)-IHPIA, has been developed for radioligand binding studies of
Ri adenosine receptors. In addition, the effects of (-)IHPIA on adenosine-mediated
responses of rat fat cells have been characterized. (-)IHPIA is slightly
less potent at Ri adenosine receptors than (-)N6-phenylisopropyladenosine
(-)PIA as assessed by adenylate cyclase and lipolysis studies.
(-)IHPIA inhibited basal adenylate cyclase activity with an IC50
of 60 nmol/l compared to an IC50 of 16.3 nmol/l for (-)PIA. (-)PIA
and (-)IHPIA inhibited adenosine deaminase-stimulated lipolysis of
intact rat fat cells with an IC50 of 0.55 and 3.6 nmol/l. The potency
of (-)N6-phydroxyphenylisopropyladenosine (-)HPIA was intermediate.
(-)HPIA has been labelled with carrier-free Na125I to very high
specific activity (2,175 Ci/mmol) and used as agonist radioligand
in binding studies of Ri adenosine receptors. The binding of (-)125IHPIA
was saturable, reversible and stereospecific. Saturation analysis
revealed two affinity states with dissociation constants (KD) of
0.7 and 7.6 nmol/l and maximal number of binding sites (Bmax) of
0.94 and 0.95 pmol/mg protein. The rate constant of association,
k1, was 3.7 X 10(8) l X mol-1 X min-1. Binding was slowly reversible
with a t1/2 of 88 min. In competition experiments specific binding
was most potently inhibited by (-)PIA, N6-cyclohexyladenosine (CHA),
(-)HPIA and (-)IHPIA, followed by 5'-N-ethylcarboxamidoadenosine
(NECA) and 2-chloroadenosine. 1,3-Diethyl-8-phenylxanthine (DPX)
and 8-phenyltheophylline were the most potent adenosine antagonists
with Ki-values of 67 and 83 nmol/l, whereas the methylxanthines 3-isobutyl-1-methylxanthine,
theophylline and caffeine had Ki-values between 1 and 21 mumol/l.(ABSTRACT
TRUNCATED AT 250 WORDS)
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