258 32 ~ Colloque SFME, Rouen-Mont St Aignan, juillet 1992

I N T E R M I T O C H O N D R I A L J U N C T I O N S IN C H E M O R E C E P T O R CELLS A N D L Y M P H O C Y T E S : U L T R A S T R U C T U R A L A N D

E X P E R I M E N T A L INVEST IGATIONS

VERNA Alain

Laboratoire de Cytologie, Universite de Bordeaux II et URA CNRS n ° 339, Avenue des Facult&s, 33405 Talence Cedex, France.

The carotid body of mammals is an arterial chemoreceptor very sensitive to hypoxia. Although mild hypoxia does not induce ultrastructural changes in the rabbit carotid body, severe hypoxia induces intermitochondrial junctions (IMJ) in chemoreceptor cells. These structures appear as septate-like junctions relating the outer membranes of mitochondria, lying very close to each other. Experimental studies, in vivo and in vitro, lead to the following conclusions : 1) IMJ appear within a few minutes after the beginning of hypoxia

(either histotoxic or anoxic) but their number does not increase with time. 2) the phenomenon is reversible: on return to normoxic conditions, IMJ disappear quickly. 3) oligomycin does not induce the formation of IMJ, in spite of ATP decrease as large as after cyanide doses sufficient to induce IMJ. 4) the formation of IMJ is inhibited by the absence of Ca 2+ in the extracellular medium. 5) the septate material is still visible after osmium-free fixation, suggesting a mostly proteic nature. 6) protein synthesis inhibition with cycloheximide or chloramphenicol (for 3 h, in vitro) does not inhibit the formation of IMJ in response to cyanide.

These results suggest that IMJ are linked to the inhibition of the respiratory chain and could be due to a redistribution of proteins which bind to mitochondrial outer membranes during hypoxia.

However, IMJ are not specific of carotid body chemoreceptor cells and may be observed in other cell types such as human lymphocytes (1). Some preliminary experiments suggest that in this cell type, IMJ are also linked to hypoxic conditions, during storing in plastic bags. The occurrence of IMJ in lymphocytes will allow to undertake biochemical investigations.

(1) RICHARD S., MASSE J.-M., GUICHARD J. et BRETON-GORlUS J. (1990). BioL Cell, 70, 27-32.

EFFECTS OF SOMAN, A N E U R O T O X I C , ON THE C E R E B R A L A S T R O C Y T E S P L A S M A M E M B R A N E S GRANGE-MESSENT Val6rie and BOUCHAUD Claude Universit~ PARIS VI, Laboratoire de Cytologie, C.N.R.S. U.R.A. 1488, 7 quai St-Bernard 75252 Paris Cedex 05

Soman, an irreversible cholinesterase inhibitor, is an epileptogen organophosphate. Very liposoluble, it crosses easily membranes and produces, among different effects, a postponed neuronal death and an astrocytic oedema. This long-duration astrocytic hypertrophy is particularly evident in the end-feet contiguous to cerebral microvessels. Effects of 0.9 LD 50 doses of soman (90 p.g/kg in the rat) administered subcutaneously, were investigated on cerebral astrocytio plasma membranes. On replicas obtained by freeze-fracture, astrocytic plasmalemmas are identifiable by the "assemblies" or "orthogonal arrays" (O.A.), i.e. square aggregates of uniform intramembranous particles (I.M.P.). These O.A. are now considered as potassium channels.

The main results of this study concern: a)- The density of O.A. which decreases in the plasmalemma contiguous to the endothelial cells as well as in the other parts of the astrocytic plasma membranes. b)- The frequent regrouping of I.MP. in "patches" and numerous cleavage planes. No other neuronal or glial plasmalemmas exhibit comparable features.

The faintest density of the O.A. could reflect the increase of the plasmalemma surface due to the astrocytic oedema. More probably, it results from a dissociation of the O. A. which are particularly labile membrane specializations(I). The astrocytic plasmalemma seems to be especially sensitive to the action of soman. Inasmuch soman increases plasmalemma fluidity(2) it could increase the lateral diffusion of the I.M.P. in the lipid bilayer.

(1) LANDIS D.M. and REESE T.S. (1981). J. Cell Biol., 88, 660-663. (2) STALC A. and al. (1989). Pharmacol. Toxicol., 64, 144-146. Supported by grant D.R.E.T. 91-156. Technical assistance o1 I. S. Delamanche and D. Raison.

DEPOSIT AND DEGRADATION OF YOLK PROTEIN IN

TROUT OOCYTES, EGGS AND EMBRYOS. AN

IMMUNOCYTOCIIEMICAL AND BIOCIIEMICAL STUDY.

~ "D2Rlean-Maric and SIRE Marie-France

UIL4 1134 CNRS, Laboratoire de Physio/ogie Cellu/aire et M6tabolique des Poissons, bat 447, UniversilE Paris-Sud, 91405 Orsay cedex.

Ill the eggs ~ most invertebrates and of a number of vertebrates, the mother "packages" the nutrients that the embryo will use for its development. In fish and amphibians, the essential process in oncyte growth is the accumulation of proteins derived from vitcllogcnin (VTG).

The present study in rainbow trout reports the sequential structural and biochemical analysis of the events leading to: 1) protcolytic cleavage of proteins taken up by the growing oocyte, in particular VTG, 2) embryonic hydrolysis of yolk proteins derived from VTG and which cover the embryo's needs during the two successive phases of its development.

It is shown that VTG is cleaved in the multivcsicular bodies formed during prcvitcllogencsis and early exogenous vitellogenesis. The action of a cathepsin D-like enzyme transforms VTG into lipovitellin and phosvitin, which are stored. Before the formation of the yolk vascular system, embryonic development occurs as a result of the degradation of yolk proteins transferred to cells during ~egmcntation. After the vascular system forms, yolk proteins are :legraded on the surface of the coalescent yolk mass of the vitelline vesicle. In this zone of lysis, protcolytic cleavage occurs via the action 3f a cathcpsin L-like enzyme synthesized by the yolk syncytium. ~nino acids are either transferred to the embryo in the blood or are ased by the syncytium to synthesize apolipoproteins.

PERINUCLEAR CYTOSKELETON AND MORPHOGENESIS OF SPERM CELLS

FOUQUET Jean-Pierre et KANN Marie-Louise.

Groupe d'Etude de la Formation du Gamete M~le

UFR Biom4dicale, 45, rue des Saints-P~res

75270 Paris Cedex 06

The perinuclear cytoskeleton of mammalian spermatids particularly the subacrosomal layer is thought to have a major role in nucleus-acrosome association and in shape changes of the head during spermiogenesis. To test these hypotheses, two models have been investigated : the experimentally cryptorchid rabbit which develops spermatids with a rudimentary and fragmented acrosome and the blind--sterile mutant mouse which produces acrosomeless spermatids. Actin (i) and calmodulin (CAM) (2), two marker components of the subacrosomal layer were surveyed by immunogold procedures. In abnormal round and elongating spermatids of rabbit end mouse models as in normal control spermatids a subacrosomal layer developed with a similar organization and actin and CaM distribution. This indicated that the subacrosomal layer forms independently of the acrosome. The timing of actin and CaM labeling during the steps of spermatid morphogenesis suggests that the subacrosomal filamentous actin might be a transitory scaffolding regulated by the Ca2+-CaM complex and involved in the assemblage of other components of the perinuclear cytoskeleton. However, this layer is not sufficient to monitor a normal apical shaping of the nucleus, rather acrosome-nucleus interactions mediated by the perinuclear layer seem necessary. The results also confirm that the microtubules of the manchette are involved in the caudal nuclear shaping.

(1) FOUQUET J.P and KANN M.L. (1992). Microsc. Res. Tech. 20, 251-252.

(2) KANN M.L., FEINBERG J., RAINTEAU D., DADOUNE J.P., WEINMAN S. and FOUQUET J.P. (1991). Anat. Rec. 230, 481-488.

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