16s Biochemical Society Transactions ( 1 995) 23
Modulation of adenosine signalling in sheep adipose tissue
by growth hormone.
ROSEMARIE DORIS, ELAINE KILGOUR, MILES D.
HOUSLAY*, GORDON E. THOMPSON and RICHARD G .
VERNON
Hannah Research Institute, Ayr, KA6 5HL and *Department of
Biochemistry, University of Glasgow, G12 8QQ
Adenosine is a paracrine/autocrine antilipolytic agent that binds
to A, receptors of adipocyte membranes, thereby activating
GTP-binding proteins, G,, which inhibit adenylate cyclase
activity and hence lipolysis.
The mechanism whereby GH facilitates lipolysis is not
fully explained, however it is possible that GH exerts its effects
by modulating the acute regulation of this process by other
hormones rather than having a direct effect on lipolysis itself.
Initially studies focused on modulation of response of
adipocytes to catecholamines, but very recent studies [ I , 21
suggest that the G,-mediated antilipolytic system is a major
target of GH action. We have investigated this further by (a)
treating sheep with recombinant bovine growth hormone,
(Monsanto) 10 mg per day for 7 days in vivo and (b) by
maintaining sheep adipose tissue explants in culture with GH
(metabolic activities and other processes are particularly well
maintained in sheep adipose tissue during culture [3]. For
culture, adipose tissue explants were preincubated for 24h with
no hormones and then GH (100ng/ml) was added for the next
24h. In both cases, response to adenosine in vitro was
determined using N6-phenylisopropyladenosine (PIA) as
described previously [2] (in essence lipolysis was activated with
100 nM isoprenaline and inhibited with various concentrations
Treatment of sheep in vivo with GH decreased
maximum inhibition of isoproterenol-stimulatedlipolysis by 100
nM PIA from 68.8 f 11.4 to 27.8 f 9.2% (P<0.05, results
means f SEM of 5 observations). This effect of GH was not
accompanied by any discernable change in the number of
adenosine receptors or amounts of the various isoforms of G,
(measured as described previously, [2]). This suggested that
a decrease in G, activity was occumng (GH also decreased
response to PGE, which activates G , but via a distinct
receptor).
Maintenance of sheep adipose tissue in culture for up to
48h in the absence of added hormones had no effect on
response to PIA. Addition of GH for 24h again decreased
(P < 0.05) maximum response to adenosine from 72.8 to 40.2%
inhibition of isoprenaline-stimulatedlipolysis (results are means
of 4 observations, SED = 9.8). Again there was no change in
either the number of adenosine receptors or amounts of G,
isoforms (not shown).
As there is evidence that G,2a can be phosphorylated by
PKC and at least one other kinase [4] we tested the effect of
protein kinase and phosphatase inhibitors in the tissue culture
system. Addition of 100 pM H7 (a protein serine kinase
inhibitor) completely abolished the effects of GH on response
to PIA (Table 1) while having no apparent effect on control
tissue. This is consistent with a kinase mediating the effects of
GH on the G, system. Further evidence for this was obtained
using the protein-serine phosphatase inhibitor, okadaic acid.
Addition of this agent to the culture medium mimicked the
effect of GH; effects of the two agents were additive (Table 1).
Table 1. Effect of H7 (100 p M ) and okadacic acid (10 nM)
on growth hormone induced inhibition of PIA action on
isoprenaline-stimulated lipolysis
Tissue was cultured for 24h without additions after which GH,
H7 and okadaic acid were added, singly or in combination, for
the next 24h. Lipolysis was measured in the presence of
isoprenaline (100 nM) and adenosine deaminase (0.8 pg/ml)
PIA (100 nM). Results are means of 4 observations; SED was
7.2 and 7.3 for the H7 and okadaic acid experiments
respectively.
% Inhibition of isoprenaline-
stimulated lipolysis
GH
Agent....
H7
Okadaic acid
-
+
-
+
25
36
44
19
11
34
23
5
These studies thus show that GH can enhance lipolysis
by decreasing the effects of agents acting via the G,-based
antilipolytic system; the effect of GH appears to involve protein
serine phosphorylation.
Lanna, D.P.D., Houseknecht, K.L., Harris, D.M., &
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Vernon, R.G. & Sasaki, S. (1991) In Physiological
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