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Functional domin in contact site a of Dictyostelium discoideum
Cell Biology
International
Reports,
Vol. 14, Abstracts
NAKLoc;I(:---mMt&EKselLci-m3islN1K~mcF.~
P593
ADULT RAT. Tcws, ROSA FENOLL,
JWEP
P594
JoRox BATLLE, PIERA, MARIANGEL LANUZA, AND DE MEDXCINA REUS. UNIVERSITAT
VERONICA FACULTAT
4320 1 REM,
Francesca Passafaro
MANEL SANTAFE, EHILI MAVAVO. DE BARCELONA.
TARRAGONA, SPAIN.
IN THE PRESENT WORK WE TRIED T O ANALIZE IF SHALL "PnvSIOLOGIC" VARIATIONS IN THE LOCP(OTOR ACTIVITY OF NORNAL ADULT RATS DURING A BRIEF PERIOD OF TIME CA,, INDUCE RENODELLING CHANGES IN THE MOTOR NERVE TERHINALS. THE POSSIBLE EXISTENCE OF PLASTIC CIIANGES IN THE BRANCHING PATTERN AND THE CONPLEXITV OF,SILVER-IMPREGNATED MOTOR NERVE TERMINALS WAS STUDIED, BY NORPHOMETRIC ANALYSIS ( 1 ) , I,, THE EXTENSOR DIGITORUM LONGUS MUSCLE OF ADULT RATS HOUSED IN METABOLIC CAGES OR TRAINED T O WALK. MORPHOHETRIC DATA INGICATE CLEARLY THAT SHALL AND EASILY REPRODUCIBLE “PHvSIoLoGIC” CHANGES IN THE LOCOHOTOR ACTIVITY OF THE NORNAL ADULT HANNALS, ARE FOLLOWED B” PRECISE NORPHOLOGTC CHANGES. SPECIFICALLY, A PRECISE INVERSE RELATION BETWEEN THE ANOUNT OF LOCONOTOR ACTIVITY AND THE COHPLEXITV FINALLY ATTAINED BY THE MOTOR NERVE ENDINGS.
Tam
J , FENOLL R, MAVAVO E, AND SANTAFA M. ( 1990 ) . BRANCHING PATTERN OF THE MOTOR NERVE ENDINGS IN A SKELETAL MUSCLE OF THE ADULT RAT. J: ANAT. l6@, 123-135.
(1)
THIS CIRIT
STUDY WAS SUPPORTED
(43-88
Swm
M),
BY GRANTS
IXICYT
FROH:
Bl8!3-W5)
PO FISSS
(83-
1645).
P595
POST-DENEWATURY
PROPRRTIlB
OF ADULT
RAT
SKRLECALWSCLEPIBRESINCULllJRE Paola
Loren~on~*~,
Fabio
Grohovaz2,
Fabio
Ruzzierl,
Robert Zorec3. 1Inst. of Physiology, Univ. of Trieste, Italy, 'CNR Ctr. Cytophanuacology 6 B. Ceccarelli Ctr., Univ. of Milan, Italy, 31nst. of Pathophysiology, Univ. of Ljubljana, Yugoslavia. We studied
culture flexor using
mammalian
without digitorus
and (jAChRs)
receptors clamp and
acetylcholine
using
patch
After
5-7
along
the entire
and kinetics denewated e jAChRs :
fibres
maintained
in
previous denervation in viva. Rat brevis muscles were dissociated
collagenase
Junctional
muscle
days
of
cultured
properties muscles
up to
two
weeks.
and extrajunctional (ejAChRs) were studied morphological techniques.
culture,
fibre
ejAChRs
membrane,
changed (jAChRs:
were
and
as 875
inserted
conductance
in the in pS, 7-2.0
vivo ms;
=48 pS, 71'1.5 ms, ~2~6.8 ms). In long term culture the open time of ACh channels was less voltage sensitive than in freshly dissociated muscles. steep
synaptic following together culture
In agreement density gradient area was
with of
in
vivo
denervation,
the
jAChRs on the border of retained in the few hours dissociation. All these results taken
suggest
acquire
that
dissociated
denervation-like
Supplement
muscle
properties.
fibres
in
1990
Specific binding
227
localization of*-Conotoxin sites at the frog aotor end plate.
Torri Tarelli, Kmanuele and Francesco Clementi.
Sher,
Maria
CNR Str of Cytopharmacology and "B. Ceccarelli" ctr , Dept. of Pharmacology, and Inst. of Human Anatomy, University of Milano, Italy. Omega-conotoxin (.&-(Xx), a novel neurotoxic peptide from the venom of Coniis geographus, irreversibly inhibits neurotransmitter release at the neuromuscular junction, probably frog in eracting with presynaptic voltage operated Ca !i+ -channels. 125 I-GCTX Binding experiments with performed on frog cutaneous pectoris muscles, revealed the presence of specific, saturable, high affinity binding sites. Nerve-muscle preparations were treated with purifiedq-CTx and the distribution of the bound toxin was investigated by indirect immunofluor;~;;~c~ t;f;hn;~;igl~mnorhodamine), using an monospecific anti-serum. Muscle fibres were consistently negative whereas specific fluorescent signal, indicative of concentrated LU-CT~ binding sites, was observed in the junctional areas. w-CTx immunoreactivity appeared to be present along the entire end plate arborization, as revealed by counterstaining with fluorescein-conjugated a-Bungarotoxin. It has been recently demonstrated that sera from patients with Lambert-Eaton myastenic syndrome (LBMS) contaipt+ autoantibodies against the ti-Clk sensitive Ca channel. A moderate, specific immunofluorescence staining of end plate regions was also detected in frog muscles incubated with LEMS sera.
P596
FUNCTIONAL IXIW.IN OF DICTYOSTELILM
IN
CX3lTACf DISCOIDELM
SITE
Motonobu Yoshida, Yoshitomi Iizuka. Research Institute of Food Science, Kinki University, Higashi-Osaka, Osaka, JAPAN. There are two kinds of cell contact in Dictvostelium discoideum: cell contact resistant to EDTA, which appears in the aggregation-competent stage, and cell contact sensitive to EDTA, which is present in the growth phase and later. It is generally thought that an integral membrane glycoprotein with an apparent molecular weight of 80,000 is involved in EDTA-resistant cell contact. It is named contact site A. The study was done to identify the functional domain of contact site A. A univalent antibody against purified contact site A inhibits EDTA-resistant cell contact. Here, we screened a Xgtll g. discoideum cDNA library with this antibody to obtain clones with cDNA coding for contact site A. The longest cDNA obtained was 2.0 kb. The cDNA clones with different coding regions for this site were ligated into the transformation-vector plasmid pA15TX, and the plasmid was used for the transformation of a mutant, HG764 that does not express the contact site A. We monitored the rate of recovery of the ability to participate in EDTA-resistant cell contact. Some transformed cells regained, so the ligated cDNA did in fact code for the functional domain of contact site A.
A
International
Reports,
Vol. 14, Abstracts
NAKLoc;I(:---mMt&EKselLci-m3islN1K~mcF.~
P593
ADULT RAT. Tcws, ROSA FENOLL,
JWEP
P594
JoRox BATLLE, PIERA, MARIANGEL LANUZA, AND DE MEDXCINA REUS. UNIVERSITAT
VERONICA FACULTAT
4320 1 REM,
Francesca Passafaro
MANEL SANTAFE, EHILI MAVAVO. DE BARCELONA.
TARRAGONA, SPAIN.
IN THE PRESENT WORK WE TRIED T O ANALIZE IF SHALL "PnvSIOLOGIC" VARIATIONS IN THE LOCP(OTOR ACTIVITY OF NORNAL ADULT RATS DURING A BRIEF PERIOD OF TIME CA,, INDUCE RENODELLING CHANGES IN THE MOTOR NERVE TERHINALS. THE POSSIBLE EXISTENCE OF PLASTIC CIIANGES IN THE BRANCHING PATTERN AND THE CONPLEXITV OF,SILVER-IMPREGNATED MOTOR NERVE TERMINALS WAS STUDIED, BY NORPHOMETRIC ANALYSIS ( 1 ) , I,, THE EXTENSOR DIGITORUM LONGUS MUSCLE OF ADULT RATS HOUSED IN METABOLIC CAGES OR TRAINED T O WALK. MORPHOHETRIC DATA INGICATE CLEARLY THAT SHALL AND EASILY REPRODUCIBLE “PHvSIoLoGIC” CHANGES IN THE LOCOHOTOR ACTIVITY OF THE NORNAL ADULT HANNALS, ARE FOLLOWED B” PRECISE NORPHOLOGTC CHANGES. SPECIFICALLY, A PRECISE INVERSE RELATION BETWEEN THE ANOUNT OF LOCONOTOR ACTIVITY AND THE COHPLEXITV FINALLY ATTAINED BY THE MOTOR NERVE ENDINGS.
Tam
J , FENOLL R, MAVAVO E, AND SANTAFA M. ( 1990 ) . BRANCHING PATTERN OF THE MOTOR NERVE ENDINGS IN A SKELETAL MUSCLE OF THE ADULT RAT. J: ANAT. l6@, 123-135.
(1)
THIS CIRIT
STUDY WAS SUPPORTED
(43-88
Swm
M),
BY GRANTS
IXICYT
FROH:
Bl8!3-W5)
PO FISSS
(83-
1645).
P595
POST-DENEWATURY
PROPRRTIlB
OF ADULT
RAT
SKRLECALWSCLEPIBRESINCULllJRE Paola
Loren~on~*~,
Fabio
Grohovaz2,
Fabio
Ruzzierl,
Robert Zorec3. 1Inst. of Physiology, Univ. of Trieste, Italy, 'CNR Ctr. Cytophanuacology 6 B. Ceccarelli Ctr., Univ. of Milan, Italy, 31nst. of Pathophysiology, Univ. of Ljubljana, Yugoslavia. We studied
culture flexor using
mammalian
without digitorus
and (jAChRs)
receptors clamp and
acetylcholine
using
patch
After
5-7
along
the entire
and kinetics denewated e jAChRs :
fibres
maintained
in
previous denervation in viva. Rat brevis muscles were dissociated
collagenase
Junctional
muscle
days
of
cultured
properties muscles
up to
two
weeks.
and extrajunctional (ejAChRs) were studied morphological techniques.
culture,
fibre
ejAChRs
membrane,
changed (jAChRs:
were
and
as 875
inserted
conductance
in the in pS, 7-2.0
vivo ms;
=48 pS, 71'1.5 ms, ~2~6.8 ms). In long term culture the open time of ACh channels was less voltage sensitive than in freshly dissociated muscles. steep
synaptic following together culture
In agreement density gradient area was
with of
in
vivo
denervation,
the
jAChRs on the border of retained in the few hours dissociation. All these results taken
suggest
acquire
that
dissociated
denervation-like
Supplement
muscle
properties.
fibres
in
1990
Specific binding
227
localization of*-Conotoxin sites at the frog aotor end plate.
Torri Tarelli, Kmanuele and Francesco Clementi.
Sher,
Maria
CNR Str of Cytopharmacology and "B. Ceccarelli" ctr , Dept. of Pharmacology, and Inst. of Human Anatomy, University of Milano, Italy. Omega-conotoxin (.&-(Xx), a novel neurotoxic peptide from the venom of Coniis geographus, irreversibly inhibits neurotransmitter release at the neuromuscular junction, probably frog in eracting with presynaptic voltage operated Ca !i+ -channels. 125 I-GCTX Binding experiments with performed on frog cutaneous pectoris muscles, revealed the presence of specific, saturable, high affinity binding sites. Nerve-muscle preparations were treated with purifiedq-CTx and the distribution of the bound toxin was investigated by indirect immunofluor;~;;~c~ t;f;hn;~;igl~mnorhodamine), using an monospecific anti-serum. Muscle fibres were consistently negative whereas specific fluorescent signal, indicative of concentrated LU-CT~ binding sites, was observed in the junctional areas. w-CTx immunoreactivity appeared to be present along the entire end plate arborization, as revealed by counterstaining with fluorescein-conjugated a-Bungarotoxin. It has been recently demonstrated that sera from patients with Lambert-Eaton myastenic syndrome (LBMS) contaipt+ autoantibodies against the ti-Clk sensitive Ca channel. A moderate, specific immunofluorescence staining of end plate regions was also detected in frog muscles incubated with LEMS sera.
P596
FUNCTIONAL IXIW.IN OF DICTYOSTELILM
IN
CX3lTACf DISCOIDELM
SITE
Motonobu Yoshida, Yoshitomi Iizuka. Research Institute of Food Science, Kinki University, Higashi-Osaka, Osaka, JAPAN. There are two kinds of cell contact in Dictvostelium discoideum: cell contact resistant to EDTA, which appears in the aggregation-competent stage, and cell contact sensitive to EDTA, which is present in the growth phase and later. It is generally thought that an integral membrane glycoprotein with an apparent molecular weight of 80,000 is involved in EDTA-resistant cell contact. It is named contact site A. The study was done to identify the functional domain of contact site A. A univalent antibody against purified contact site A inhibits EDTA-resistant cell contact. Here, we screened a Xgtll g. discoideum cDNA library with this antibody to obtain clones with cDNA coding for contact site A. The longest cDNA obtained was 2.0 kb. The cDNA clones with different coding regions for this site were ligated into the transformation-vector plasmid pA15TX, and the plasmid was used for the transformation of a mutant, HG764 that does not express the contact site A. We monitored the rate of recovery of the ability to participate in EDTA-resistant cell contact. Some transformed cells regained, so the ligated cDNA did in fact code for the functional domain of contact site A.
A