No description available.

The process of normal spermatogenesis was investigated in six species of the genus Dermestes (D. haemorrhoidalis, D. ater, D, peruvianus, D. lardarius, p. frischii, D. maculatus and D. vulpinus). In all of the species germ cells are connected with each other by cytoplasmic bridges and surrounded by an envelope consisting of one or more cyst cells. Therefore the time of isolation of the new predefinitive spermatogonia (A0) can be determined directly by counting the number of cells in individual cysts. Once differentiation begins, the spermatogonia undergo a series of more or less synchronous mitoses during which they change progressiveiy in important characteristics: their size diminishes and their number per cyst (i.e. 2n) increases. These changes are in the direction of, and preparatory to, meiosis. The number of generations of secondary spermatogonia is 7 in D. haemorrhoidalis, D. ater, D. peruvianus and D. lardarius, 6 in D. frischii, D. vulpinus and D. maculatus. In D. maculatus the duration of the mitotic cycle almost always shows no change in the duration of S- or G2- through 6 generations of spermatogonia with a steady cell cycle time of 20 hrs. The primary spermatocyte, while its nucleus undergoes the conspicuous changess of meiosis, characteristically grows to about 11 fold its starting volume in D. haemorrhoidalis, 8 fold in D. star, 5 fold in D. peruvianus, 9 fold in D. lardarius, 11 fold in D. frischii, 7 fold in D. vulpinus and 6 fold in D. maculatus. No typical leptotene, zygotene, pachytene or diplotene stages are visible because the euchromatic regions remain diffuse until diakinesis and diakinesis is of short duration in Dermestes. The duration of the meiotic cycle in Dermestes is correlated weakly with DNA content and varies from 12.75 days to 19 days for the period pre-meiotic S-phase to elongate spermatid. In Dermestes a certain number of germ cells die. The great majority of cell deaths occur at definitive periods of development of the spermatocytes. As far as the responses of spermatogonial and spermatocyte cells of Dermestes to x-irradiation are concerned it has been shown that: a) The last two generations of spermatogonia (In-type & B-type) appear to be the most sensitive to cell death; b) Damaged spermatogonia of all types degenerate either in interphase or upon entrance into prophase of their first post-irradiation division. The commonly-observed lack of metaphase is obviously accounted for by the degeneration in earlier stages; c) Repopulation of any follicle depends on the number of type AO spermatogonia present along the germarium membrane after irradiation; d) Spermatocytes are more sensitive to the induction of cell death at early prophase-I rather than late prophase-I; e) The distinct phenomenon of "stickiness" of the chromosomes appear to be due to a generalised physiological process; f) On the basis of dose-response kinetics, there is a variation between species with D. haemorrhoidalia being the most resistant and D. peruvianus and D. lardarius the most sensitive to x-irradiation induced spermatocyte cells death; g). On the basis of dose-response curves, A0 spermatogoniam appeared to be more resistant to x-rays induced cell death in D. haemorrhoidalis and more sensitive to x-rays induced cell death in D. lardarius; h) Distribution of cell-number in spermatocyte cysts which surviv ex-irradiation shows that a fixed proportion of cells may be lost but then the whole cyst dies. Cells may still die in small groups before this catastrophe occurs; i) There is a significant correlation between high exposure (i.e. exposure, giving 10% survival) and spermatocyte nuclear volume. The model for maintenance of a continuing supply of spermatogonia during adult life suggests that there is a group of AO spermatogonia in each follicle which divide quasidichotomously with one becoming a new A0 spermatogonia (responsible for the next generation of germ cells) and the other (the definitive spermatogonium) destined, through multiplication and differentiation, to give rise to the mature spermatozoa. The model proposed here for Dermestes species (Coleoptera) is in good agreement with other models, suggested for different orders of Insecta. The numbers of A0 spermatogonia in each follicle are estimated for each Dermestes species studied. The likelihood of sterilization of a male Dermestes depends upon three factors: i) The number of follicles per testis; ii) The number of AD cells per follicle; iii) The inherent sensititity of the AD cells.

The fine structure of the median i n direct dorsal longitudinal flight muscles has been examined throughout the first week of adult life. During this period, muscle colour changed from white to reddishbrown and the banding pattern characteristic of mature adult flight muscle was established. Associated with these changes there was an increase in myofbrilsize and the mean number of myosin filaments per myofibril,- no significant change was observed in the actin : myosin ratio. There were indications that the number of myofibrils per muscle fibre increase by " longitudinal splitting" of existing myofibrils in the first four days of adult life. A marked increase in mitochondrial size and complexity was noted with increasing age. In addition, total mitochondrial protein increased approximately 10-fold between the 9 th day of 5th instar and the 6th day of adult life. However, the increase in mitochondrial size is probably due to both the syn thesis of new mitochondrial protein and the fusion of adjacent mitochondria. The mitochondria gradually become arranged in straight columns between the myofibrils by the 5th day of adult life. The relative volume of the sarcoplasmic reticulum (SR) and T-system decreased with age. This was associated with the formation of dyadic junctions and the separation of adjacent myofibrils by sheet(s) of SR. In contrast to the situation, observed in the first few days of adult life, where dyadic junctions are situated near the Z-bands or at an oblique angle to the adjacent m of brils, in more developed flight muscle they are situated in the region of the A-bands and run parallel to the myofibrils. The distribution of the SR and T-system was affected by the penetration of tracheoles into the muscle fibres. Muscle tracheation was more-or-less fully developed within the 3rd-day of adult life. The relative volume of the tracheoles decreased with age. The physiological implications of these developmental changes in fine structure are discussed. Mitochondrial phospholipids contained five mean fatty acids at all ages studied; palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1), linoleic acid (18:2) and linolenic acid (18:3). The relative amount of stearic acid (18:0) and the ratio of unsaturated : saturated fatty acids decreased over the period between the 9th-day of the 5th instar and the 15th day of adult life. Providing a suitaible reaction medium was used, oxidative phosphorylation was demonstrable at all ages studied with the following substrates: ɑ-glycerophosphate, pyruvate plus proline and glutamate, but not with succinate. Allosteric activation of ɑ-giycerophosphate dehydrogenase by Ca(^2+) was demonstrable at all ages studied. No such activation was observed when Mg(^2+) replaced Ca(^2+) a-Glycerophosphate dehydrogenase activity increased with age. The properties of SR-ATPase (total ATPase) of Locusta flightmuscle were similar to those reported for vertebrates skeletal muscle SR-ATPase. Total ATPase activity required Mg(^2+) and was stimulated byCa(^2+). Optimal concentrations of Ca(^2+) and Mg(^2+) for total ATPase were 3.19 X 10(^-6)M (free Ca(^2+)) and 1.5 - 3mM, respectively. The pH optimum was ca. 7.6 for total ATPase activity. The Ca(^2+) -stimulated component of ATPase activity showed similar optima to total ATPase activity. In the presence of Ca(^2+) the apparent Km for ATP was decreased from 0.643mM (Mg(^2+) present) to 0.420mM. There was an approximately 4-fold increase in SR protein per thorax and in the specific activity of total and Ca(2+)- stimulated ATPase activity. The developmental changes referred to above are discussed in relation to the improved flight performance observed during the first week of adult life in Locusta migratoria L.

This thesis initially proposes an hypothesis which relates clinical findings of abnormal body monovalent cation distribution and adrenocorti-costeroid secretion in subjects suffering affective disorders to the regulation of plasma membrane cation permeability. The hypothesis is partially examined by direct investigation of the sodium and potassium content and membrane magnesium-dependent adenosinetriphosphatase activities of erythrocytes from normal and affectively disordered human subjects. Thus, whilst erythrocyte sodium content remains normal, potassium content and monovalent cation-stimulated, magnesium-dependent adenosinetriphos-phatase activity are elevated in manic depression. Further investigations are concerned with determining the influence of adrenocorticosteroids upon these and related factors in rat skeletal muscle, kidney, erythrocytes and brain. They reveal that, whilst mono valent cation-stimulated, magnesium-dependent adenosinetriphosphatase activity remains uninfluenced by adrenocorticosteroid depletion in rat brain, it is reduced in skeletal muscle and kidney, and elevated in erythrocytes. Despite evidence for an associated significant decrease in the sodium content of rat erythrocytes, the rate of sodium efflux is reduced, and the activity of another membrane-associated enzyme (acetylcholinesterase) remains uninfluenced. Although the dietary administration of isotonic sodium chloride solution is shown to counteract a number of the effects of adreno corticosteroid depletion upon rat tissues in a manner which suggests that renal sodium reabsorption is the principal hormone-sensitive target process, certain results obtained in skeletal muscle and erythrocyte studies are interpreted in terms of an extra-renal regulatory influence of adrenocorti costeroids upon plasma membrane characteristics. Experimental observations are discussed throughout in association with those established by previous workers, and in connexion with the extent to which adrenocorticosteroidal control of monovalent cation distribution may be implicated in the aetiology of manic-depression. This finally leads to a reconsideration of the clinical evidence for abnormal monovalent cation distribution in mania and depression, and of the hypothesis initially proposed.

No description available.

Several species of Actinomyces viscosus were examined using the scanning electron microscope and considerable morphological heterogeneity was seen and one species, Actinomyces viscosus WVU 398B, was observed to undergo a form of differentiation, for which the term 'vacuolation' has been introduced. This form of differentiation can be regulated by inhibitors of peptidyl transferase or translocase. Further examples of the regulation of differentiation were provided by an examination of the interaction of antibiotics and antimetabolites with a species of Bacillus cereus isolated from laboratory dust. 3-chloropropane-1-2-diol exhibited an ability to impair cell division in Staphylococcus aureus and Escherichia coli and this action may be due to the glycerol antagonism of 3-chloropropane-1,2-diol.

No description available.

Optimisation of the experimental technique (BrdU-IddU double-staining) and development of new computational tools have allowed, for the first time, a comprehensive spatio-temporal map of quantitative cell cycle times in the early vertebrate limb. A key question of limb morphogenesis is how genes create the digit pattern. An example of such a gene is Sox9, which is an early marker of chondrogenesis and is, therefore, assumed to follow a pattern similar to early stages of digit patterning. Classical chondrogenic experiments, suggest digital regions are patterned by the intermediate formation of a “digital arch” from which the digits arise in a posterior to anterior order. In contrast, a through analysis of a large number of Sox9 <i>in situs</i> revealed digital regions 1, 2 and 3 branch from a region reminiscent of the tibia (anterior zeugopod) and digital regions 4 and 5 branch from a fibula-like region (posterior zeugopod). Moreover, the Sox9 pattern first arises in digital regions 2, 3 and 4, followed by digital regions 5 and 1. The Sox9 <i>in situ</i> analysis was achieved using newly developed software for the 3D analysis of optical projection tomographic (OPT) images at a very high spatial resolution. These studies have highlighted the importance of integrating practical and computational tools in order to close the gaps in our knowledge and understanding of limb development, and developmental processes as a whole. The computational tools generated for the proliferation studies are valuable in offering a thorough means of analysis of cell cycle times and the new OPT software will be invaluable for the study of both weak and strong gene expression patterns in whole embryos. In the future, the proliferation data and 3D Sox9 <i>in situ</i> data can be incorporated into simulation software, the results of which should shed light upon the interactive effects of different factors upon the process of limb morphogenesis.

No description available.

The aims of this study were i) to characterise differences in the spatial and temporal expression of Pax6 isoforms during murine embryonic development ii) to analyse the expression of Pax6 isoforms in various <i>Pax6 </i>mutant mice during neurogenesis, and iii) to examine the effects of over-expression of the best understood isoforms, in a cell culture system. The overall aim was to elucidate the independent roles of Pax6 isoforms in organogenesis of the central nervous system. RNase protection assay was used to determine the ratio between <i>Pax6 and Pax6+5a </i>transcripts in a number of tissues of the central nervous system during neurogenesis. In most tissues studied, <i>Pax6 </i>is much more prevalent than <i>Pax6+5a </i>at embryonic day 12.5, but the ratio has fallen by embryonic day 18.5. This may be indicative of a change in the role of Pax6, from controlling proliferation to controlling neuronal differentiation. Pax6 protein expression was analysed in mice with 0, 1, 2, 8 and 14 functional copies of <i>Pax6, </i>in order to compare the levels of Pax6 expression between genotypes, and to determine if differential expression of one or more isoforms could be responsible for the mutant phenotypes. Most isoforms are down-regulated in the <i>Pax6^Sey/+ </i>eye, whilst their relative expression is more varied in the <i>Pax6^Sey/+ </i>brain. Most isoforms are significantly up-regulated in the brain and eye of mice with 8 or 14 copies of <i>Pax6, </i>but there are no differences between expression levels in the brain of the two genotypes, indicating that Pax6 is subject to autoregulation. Some Pax6 isoforms are observed in the brain of mice thought to lack functional <i>Pax6.</i>