MONDAY 30 M A R C H
POSTERSRELATEDTOS2
Mon-S2- 16
Mon-S2-17
INHIBITION OF CALMODULIN-ACTIVATED CYCLIC
NUCLEOTIDE PHOSPHODIESTERASE BY TRICYCLIC DRUGS.
C. H. Reynolds, Biochemical Research Dept.,
Wellcome Foundation Ltd., Dartford,DAl 5AH,U.K.
A number of t r i c y c l i c psychotropic drugs, e.g.
t r i f luoperazine (TFP) , chlorpromazine (CPZ) and
a m i t r i p t y l i n e ( A T ) , bind t o calmodulin (CDR) and
i n h i b i t Ca++/CDR-activated phosphodiesterase (PDE)
and most o t h e r CDR-activated enzymes. The a c t i v a t i o n of guinea-pig h e a r t PDE has been measured
with and without t r i c y c l i c s over a range of CDR
c o n c e n t r a t i o n s . A l l of t h e compounds used (TFP,
CPZ, AT, imipramine and desipramine) antagonised
t h e a c t i v a t i o n , b u t t h i s i n h i b i t i o n was overcome
a t high concentrations of CDR. With one compound
(AT) t h e apparent d i s s o c i a t i o n - c o n s t a n t f o r t h e
drug was i n v e s t i g a t e d a s a f u n c t i o n of A T conc e n t r a t i o n and found t o decrease a s t h e drug
c o n c e n t r a t i o n was increased.
T h i s suggests t h a t
CDR can be i n h i b i t e d by t h e binding of two o r
more molecules cf t r i c y c l i c compounds, i n accordance with d i r e c t binding d a t a .
Ca2+ dependent chemiluminescence i n r a t polymorphs
induced by complement a c t i v a t i o n
H a l l e t t , M.B., Luzio, J . P . X and Campbell, A . K . ,
Department o f Medical Biochemistry, Welsh National
School o f Medicine, Heath Park, C a r d i f f , U.K. and
*Department of C l i n i c a l Biochemistry, Addenbrooke's
H o s p i t a l , Cambridge, U.K.
I t i s proposed t h a t a c t i v a t i o n of complement
r e s u l t s i n a rapid increase i n i n t r a c e l l u l a r free
Ca2+.
This can occur before c e l l l y s i s and a l s o
p l a y a key p a r t i n non-lytic e f f e c t s of complement
on c e l l s .
The increase i n i n t r a c e l l u l a r Ca2+ i s
caused e i t h e r by i n t e r a c t i o n of c e l l s u r f a c e
r e c e p t o r s with complement components o r products
such as C3a and C 5 c a l t e r n a t i v e l y by t h e
terminal complex C56789.
The l a t t e r e f f e c t has
been demonstrated d i r e c t l y i n e r y t h r o c y t e 'ghosts'
using t h e photoprotein o b e l i n a s an i n d i c a t o r of
This hypothesis has
i n t r a c e l l u l a r f r e e Ca2+.
been t e s t e d by studying t h e e f f e c t of complement
a c t i v a t i o n on t h e Ca2+ dependent chemiluminescence
response o f polymorphs p o t e n t i a t e d by l u m i n o l .
Mon-S2-18
Mon-S2-19
ETHACRYNIC ACID (ECA) AND a-ADRENERGIC
CONTROL OF HEPATIC GLYCOGENOLYSIS.
A. Jakob, N. Isler & S. Diem,
Dept. of Biochemistry, University,
CH-4051 Basel, Switzerland.
a-Adrenergic activation of hepatic glycogenolysis was previously shown to be parralleled by a release of Ca2+ and a redistribution of K+. We used ECA to further
investigate the relationship between active ion transport and a-adrenergic responses of perfused livers. ECA enhanced
glycogenolysis, produced a massive release of X+ and a moderate Ca2+ l o s s . It
suppressed the responses to phenylephrine
(0.5 pM). In purified liver plasma membranes (Na+-K+)ATPase and ouabain-insensitive ATPase activities were inhibited
and binding of 3H-epinephrine to a-adrenergic sites was decreased by ECA. The
latter observation may explain the loss
of responsiveness to phenylephrine.
R 24571, A N E W AND VERY POTENT I N H I B I T O R OF
CALVODULIN ACTIVATION OF PDE
H. Van B e l l e , Janssen Pharmaceutica Research Labo-
of the CDR-activation of b r a i n PDE. The compound
has been compared w i t h t r i f l u o p e r a z i n e ( t h e r e f e rence compound used i n many s t u d i e s on CDR) and
with p e n f l u r i d o l ( t h e most p o t e n t i n h i b i t o r desc r i b e d s o f a r - Levin, Weiss 1 9 7 9 ) . The 1 5 0 values
on Ca2+-CDR-PDE a r e : t r i f l u o p e r a z i n e : 2.6 X 10-6M
p e n f l u r i d o l : 3.7 X 10-7M and R 24571 : 5 X IO-'M.
The compound a l s o s t r o n g l y i n h i b i t s the CDR-dependent Ca2+-ATPase from e r y t h r o c y t e s . Preliminary
d a t a on t h e e f f e c t of R 24571 on o t h e r enzymes
w i l l be r e p o r t e d . R 24571 i s not t o x i c t o mice embryonal c e l l s i n c u l t u r e a t concentrations < 1.5 X
lod6# nor a t 6 10 mg/kg i . p . i n mice o r r a t s . Its
high potency may open new p e r s p e c t i v e s f o r s t u d i e s
on the involvement of calmodulin i n b i o l o g i c a l
processes.
Mon-S2-20
Mon-S2-21
ANALYSIS BY ISOELECTRIC FOCUSING AND ELECTROPHORESIS OF THE MICROSOMAL PROTEINS OF SKELETAL
MUSCLE.
U n i d i r e c t i o n a l c a l c i u m uptake a t maternal and
f e t a l surfaces o f t h e i s o l a t e d d u a l l y - p e r f u s e d
guinea-pig placenta.
J.H. Sweiry, B.M. Eaton, G.E. Mann&D.L.Yudilevich
Dept o f Physiology, Queen E l i z a b e t h College, Univ.
of London, London, W8 7AH, U . K .
.
r a t o r i e s , B-2340 Beerse, Belgium.
R 24571, an analogue of t h e a n t i s e p t i c sepazo-
n i u m , has been found t o be a very p o t e n t i n h i b i t o r
M. Lynn O'Connor and Denis R. Headon, Department
o f Biochemistry, U n i v e r s i t y College, Galway.
I r e 1and.
Uptake i n t o t h e t r o p h o b l a s t from maternal and f e t a l
c i r c u l a t i o n s was s t u d i e d u s i n g a p a i r e d - t r a c e r
d i l u t i o n technique.
An i n t r a - a r t e r i a l i n j e c t i o n
o f 45Ca and "Na ( e x t r a c e l l u l a r r e f e r e n c e ) was
f o l l o w e d by r a i d sampling from b o t h venous e f f l u ents. Maximal '%a uptake (Umax) was determined
from (1-45Ca/22Na) and an assymetry o f uptake was
observed. A t 0.5 mM Ca2+, Um on t h e f e t a l s i d e
was 58%f 6, ( n = 3 placentae? and f o l l o w e d by
r a p i d t r a c e r e f f l u x . Umax on t h e maternal s i d e was
19% t 3 ( n = 4 ) and remained constant. Umax was r e duced by i n c r e a s i n g t h e p e r f u s a t e c o n c e n t r a t i o n o f
Ca2+ and i n f l u x was s a t u r a b l e on b o t h s i d e s o f t h e
trophoblast.
(Funded b y ARC)
Microsomal p r o t e i n s from r a b b i t s k e l e t a l muscle
have been we1 1 c h a r a c t e r i sed by p o l y a c r y l amide
gel e l e c t r o p h o r e t i c techniques y i e l d i n g a
r e l a t i v e l y simple p r o t e i n composition p r o f i l e .
However when two dimensional separations a r e
c a r r i e d o u t using i s o e l e c t r i c focusing i n the
f i r s t dimension and g r a d i e n t p o l y a c r y l ami de g e l
e l e c t r o p h o r e s i s i n t h e second dimension, a n a l y s i s
by such techniques i n d i c a t e s t h a t t h e r e i s no
s i n g l e major p r o t e i n band accounting f o r 70-80%
o f t h e t o t a l p r o t e i n . Large d i f f e r e n c e s i n
p r o t e i n p a t t e r n s a r e observed between heavy and
l i g h t membranes i s o l a t e d by sucrose d e n s i t y
g r a d i e n t c e n t r i f u g a t i o n when analysed by t h i s
technique.
')Biochim. Biophys. Acta 596, 315-319 (1980).
')Placenta (Suppl.), i n press.
133P