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Modulation of cardiac L‐type Ca 2+ channels by GTPγS in response to isoprenaline, forskolin and photoreleased nucleotides
Author(s) -
Kozlowski R.Z.,
Goodstadt L.J.,
Twist V.W.,
Powell T.
Publication year - 1994
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.1476-5381.1994.tb14052.x
Subject(s) - isoprenaline , forskolin , gtp' , nucleotide , chemistry , cyclic gmp , biophysics , cyclic nucleotide , modulation (music) , endocrinology , medicine , biochemistry , biology , enzyme , physics , stimulation , receptor , gene , acoustics
1 Using the patch‐clamp recording technique, we have investigated the effects of chronic intracellular application of guanosine thiotriphosphate (GTPγS) by cell dialysis, on the potentiation of L‐type Ca 2+ currents ( I Ca ) by isoprenaline and forskolin and also by GTPγS and cyclic AMP released intracellularly by flash‐photolysis of their caged derivatives. 2 GTPγS prevented enhancement of I Ca by isoprenaline with an IC 50 of ∼ 10 μ m and considerably reduced the ability of forskolin to increase I Ca . In addition GTPγS also reduced the time‐to‐peak response for potentiation of I Ca by forskolin. Responses to forskolin were abolished by co‐dialysis of cells with the cyclic AMP antagonist, R p ‐adenosine‐3′‐5′‐mono‐thionophosphate (R p ‐cAMPS). 3 Photoreleased GTPγS (PR‐GTPγS; ∼23 μ m ) generally induced a biphasic increase in I Ca . This response was also inhibited by chronic intracellular dialysis with GTPγS with an IC 50 of ∼1 μ m . 4 Pretreatment of cells with pertussis toxin (PTX) reversed the inhibitory effect of 100 μ m GTPγS on isoprenaline‐induced stimulation of I Ca . However, PTX pretreatment did not restore the activating action of PR‐GTPγS inhibited by chronic application of GTPγS. 5 Photoreleased cyclic AMP (∼5 μ m ; PR‐cyclic AMP) increased peak I Ca . This effect was inhibited by dialysis of cells with R p ‐cAMPS and by stimulation of I Ca by the phosphodiesterase inhibitor 3‐isobutyl‐1‐methylxanthine. Co‐dialysis of cells with uncaged GTPγS reduced the time‐to‐peak for PR‐cyclic AMP mediated activation of I Ca but did not affect the magnitude of the response. 6 It is concluded that chronically applied GTPγS can (i) inhibit activation of I Ca by isoprenaline by interacting with a PTX‐sensitive guanosine nucleotide binding (G‐) protein located upstream of adenylate cyclase (possibly G i ) and (ii) accelerate the response to cyclic AMP dependent phosphorylation possibly by interacting with a G‐protein coupled directly to the channel. 7 In view of this diverse range of effects, care should be taken when using GTPγS to characterize G‐protein‐mediated events, since the resulting physiological response may be due to activation of several G‐protein containing pathways.