THE SPERM CENTRIOLE PERSISTS DURING EARLY EGG CLEAVAGE IN THE INSECT CHRYSOPA CARNEA (NEUROPTERA, CHRYSOPIDAE)

Transcription

1 J. Cell Sci. 42, (1980) 221 Printed in Great Britain Company of Biologists Limited 1980 THE SPERM CENTRIOLE PERSISTS DURING EARLY EGG CLEAVAGE IN THE INSECT CHRYSOPA CARNEA (NEUROPTERA, CHRYSOPIDAE) MICHAEL FRIEDLANDER Department of Biology, Ben Gurion University of the Negev, Beer Sheva, Israel SUMMARY The giant centriole of the sperm of Chrsyopa carnea persists in the fertilized egg. The giant centriole appears together with a regular small centriole at one pole of the first cleavage spindle; two small centrioles are found at the other pole. During the subsequent divisions, the giant centriole appears either at one pole of one of the spindles, as in the first cleavage, or close to one interkinetic nucleus. All the other centrioles are regular in size. Therefore, the assumption that the mature spermatozoa of insects contain no true centrioles is not valid, at least for Chrysopa. INTRODUCTION The question of whether the centrioles of insect spermatozoa continue as distinct morphological and functional entities throughout spermatogenesis and embryogenesis is still subject to dispute. Basing their conclusions on essentially the same ultrastructural data on sperm differentiation, some authors have indicated that one centriole is generally found in mature insect spermatozoa (Friedlander & Wahrman, 1971; Friedlander & Gitay, 1972), while others have concluded that the centrioles degenerate and disappear during spermatogenesis (Phillips, 1970; Baccetti & Afzelius, 1976). Early studies (Huettner, 1933) on the presence of the sperm centriole in the cleaving egg are inconclusive as the tiny sperm centriole could not be definitely distinguished with the light microscope from the other egg centrioles having the same dimensions. Systematic ultrastructural analyses of this nature have not been made, probably due to the difficulty of finding, recognizing and sectioning the sperm centriole in the bulky egg- In the present study we took advantage of the giant centriole (up to 8 /tm) ofneuropteran spermatozoa (Friedlander & Wahrman, 1966) to follow with the light microscope the fate of the sperm centriole in the cleaving egg. These giant centrioles are about 15 times larger than the other centrioles of the eggs (e.g. Mahowald, 1963) and could be identified with certainty.

2 M. Friedldnder MATERIALS AND METHODS Chrysopa carnea (Neuroptera, Chrysopidae) was used as it has the longest sperm centriole known among insects (Friedlander & Wahrman, 1966). Eggs were collected at laying or after that, every 5-16 min for 2 h, then every 20 min until 5 h after laying. At laying, the oocytes are generally at the first meiotic prophase but a few are at the first anaphase-telophase or even at the second meiotic metaphase. Five to ten eggs were fixed each time. For fixation, the eggs were immersed in Flemming's strong bulkfluid (Darlington & LaCour, 1962) and immediately punctured with a thin tungsten needle to facilitate penetration of the fixative. Testes of adults werefixedin the same solution. Parafin sections, 10 /tm thick, of both eggs and testes were stained with crystal violet (Soumalainen, 1952). RESULTS Centrioles during meiosis There are 2 giant centrioles at each spindle pole of the male first meiotic division (Fig. 1) but only one centriole is found ateach pole of the second division. The spermatids have one giant centriole (Fig. 2) from which the axoneme of the flagellum develops. For further details see Friedlander & Wahrman (1966) and White (1973). Centrioles and asters are absent throughout the meiotic divisions of the oocytes (Fig. 3). No separate polar bodies are formed as the chromosomes reaching the outer poles at both meiotic divisions remain in the periplasm, beneath the surface of the oocyte. Fig. 1. Section of a first spermatocyte of Chrysopa carnea at metaphase. Two giant centrioles forming an angle of about 170 are seen at the upper pole; only part of these centrioles are in the focal plane of the picture. A transverse section of a centriole is shown at the lower pole. The X and Y sex chromosomes are close to the poles forming the socalled distance pairing (White, 1973) x Fig. 2. Section of an early spermatid of Chrysopa carnea showing the giant centriole. 11, nucleus, x Fig. 3. Section through the spindle of the first meiotic anaphase-telophase of the oocyte of Chryospa carnea. Centrioles are absent from the spindle poles, x 1800.

3 Sperm centriole in the Chrysopa egg 223 Evolution of the centriole of the fertilizing spermatozoon The spermatozoon enters the egg through the micropyle which is then immediately closed by the operculum. Both the head and flagellum undergo profound changes during the development of the oocyte nucleus into the female pronucleus but only data concerning the sperm centriole are included here. During telophase of the first oocyte division, the flagellum disconnects from the sperm nucleus but remains close to it; somewhat later the giant centriole separates from the axoneme. The giant centriole is surrounded by a conspicuous aster (Fig. 4). The development of the sperm nucleus continues until the spherical shape of the male pronucleus is achieved, which coincides with the formation of the female pronucleus. The giant centriole, surrounded by its aster, accompanies the male pronucleus (Fig. 5) until the fusion of the pronuclei and the formation of the first cleavage spindle. There is only one giant centriole in the early embryo; all the other centrioles are of the regular short type. The giant sperm centriole appears together with a regular small centriole at one pole of the first cleavage spindle; the other pole contains 2 small centrioles. During the successive mitotic divisions the giant centriole appears either at one pole of one spindle, as at the first cleavage spindle, or close to one interkinetic nucleus. This regular behaviour of the giant centriole continues during the peripheral migration of the cleavage nuclei (Figs. 6, 7). Later stages of embryogenesis were not studied. DISCUSSION The sperm centriole does not disappear The giant centriole of the spermatozoon of Chrysopa cornea persists. In the fertilized egg, this centriole appears at the poles of the mitotic spindles and is transmitted apparently unchanged and in an orderly fashion throughout the cleavage divisions. Therefore, the assumption that the mature spermatozoon of insects contains no true centrioles (Baccetti, 1972) is not valid, at least for Chrysopa. This idea was based mainly on the fact that an extremely modified centriole, lacking the customary pattern of 9 triplets of microtubules, is found in the mature spermatozoa of different insect species (e.g. Szollozi, 1975), after the role of this centriole in the morphogenesis of the flagellum has ended (e.g. Fawcett, 1972). Indeed, centrioles have been observed to disassemble in fully differentiated cells of metazoans such as pinealocytes (Lin, 1970) and Sertoli cells (Friedlander, 1972). Moreover, it appears that the centrioles of mammalian spermatozoa also disassemble (Fawcett, 1972) and centrioles are clearly absent from the early cleavage stages of mammalian eggs (Szollozi, 1972). However, the ultrastructural modifications which have been found in insect centrioles do not necessarily indicate that these are degenerating centrioles (Friedlander & Wahrman, 1970). There are numerous cases of centrioles lacking the customary patterns but acting normally during spermatogenesis in flagella genesis and cell division. The most well known among these are the centrioles having up to 350 singlets which develop during the spermatogenesis of fungus gnats

4 224 M. Friedldnder

5 Sperm centriole in the Chrysopa egg 225 (Phillips, 1966, 1967; Shay, 1973). Therefore, the definition demanding the presence of 9 triplets of microtubules is useful only for centrioles having this pattern. To recognize other centrioles, a less morphologically restricted but more functional definition has previously been proposed (Friedlander & Wahrman, 1971): Centrioles are organelles composed of groups of parallel microtubules (not only those having 9 triplets), acting actually or potentially as basal bodies; in metazoan cells, the centrioles appear generally (but not always) at the spindle poles during division. According to these considerations and to the findings of the present report, it should be stressed that the mature spermatozoa of Chrysopa carnea, and probably of other insect species as well, do contain centrioles. REFERENCES BACCETTI, B. (1972). Insect sperm cells Adv. Insect Physiol. 9, BACCETTI, B. & AFZELIUS, B. A. (1976). The biology of the sperm cell. Monogr. dev. Bio]. 10, DARLINGTON, C. D. & LACOUR, L. F. (1962). The Handling of Chromosomes. London: Allen & Unwin. FAWCETT, D. W. (1972). Observations on cell differentiation and organelle continuity in spermatogenesis. In The genetics of the Spermatozoon, (ed. A. Beatty & S. Gluecksohn-Waelsh), pp Edinburgh & New York: Departments of Genetics of the University of Edinburgh and of the Albert Einstein College of Medicine. FRIEDLANDER, M. (1972). Transient centrioles in Sertoli cells of the mole-rat Spalax ehrenbergi. Cytobios 6, FRIEDLANDER, M. & GITAY, H. (1972). The fate of the normal-anucleated spermatozoa in inseminated females of the silkwork Bombyx mori. J. Morph. 138, FRIEDLANDER, M. & WAHRMAN, J. (1966). Giant centrioles in neuropteran meiosis.^. Cell Sci. 1, FRIEDLANDER, M. & WAHRMAN, J. (1970). The spindle as a basal body distributor. A study in the meiosis of the male silkworm moth (Bombyx mori). J. Cell Sci. 7, FRIEDLANDER, M. & WAHRMAN, J. (1971). The number of centrioles in insect sperm. A study in two kinds of differentiating silkworm spermatids. J. Morph. 134, HUETTNER, A. K. (1933). Continuity of the centriole in Drosophila melanogaster. Z. Zellforsch. mikrosk. Anat. 19, Lin, H. S. (1970). The fine structure and transformation of centrioles in the rat pinealocyte. Cytobios 6, MAHOWALD, A. P. (1963). Ultrastructural differentiations during formation of the blastoderm in the Drosophila melanogaster embryo. Devi Biol. 8, PHILLIPS, D. M. (1966). Observations on spermiogenesis in the fungus gnat Sciara coprophila. J. Cell Biol. 30, PHILLIPS, D. M. (1967). Giant centriole formation in Sciara. J. Cell Biol. 33, Figs Sections of fertilized eggs of Chrysopa carnea. Fig. 4. The giant centriole of the sperm is separated from the flagellum and surrounded by a conspicuous aster, x Fig. 5. The 2 pronuclei. The giant centriole of the sperm is close to the male pronucleus. x Fig. 6. A stage in the formation of the syncytial blastoderm showing the nuclei still distant from the egg surface (m). x 850. Fig. 7. Two nuclei as in Fig. 6. The giant sperm centriole (arrow) is adjacent to the left-hand nucleus in the figure, x 2400.

6 226 M. Friedldnder PHILLIPS, D. M. (1970). Insect sperm: Their structure and morphogenesis. J. Cell Biol. 44, SHAY, J. W. (1973). Ultrastructural observations on spermiogenesis in the fungus gnat Rhynchosciara sp. In The Functional Anatomy of the Spermatozoon (ed. B. A. Afzelius), pp Oxford and New York: Pergamon. SUOMALAINEN, H. O. T. (1952). An improved method for the use of crystal violet and allied dyes as chromosome stains. Soumal. ela'in-ja kasvit. Seur. van. Tiedon. Poyta'k. 7, SZOLLOZI, A. (1975). Electron microscope study of spermiogenesis in Locusta migratoria (Insecta Orthoptera). J. Ultrastruct. Res. 50, SZOLLOZI, D. (1972). Changes of some cell organelles during oogenesis in mammals. In Oogenesis (ed. J. D. Biggers & A. W. Schuetz), pp London: Butterworth. WHITE, M. J. D. (1973). Animal Cytology and Evolution. Cambridge University Press. (Received 20 July 1979)