HISTOCHEMICAL AND ELECTRON MICROSCOPIC OBSERVATIONS ON THE STEROID HORMONE-SECRETING CELLS IN THE TESTIS OF THE JAPANESE RED-BELLIED NEWT, CYNOPS PYRRHOGASTER PYRRHOGASTER

The testis of the Japanese red-bellied newt, Cynops pyrrhogaster pyrrhogaster, caught in the middle of May was investigated in order to demonstrate the presence of steroid hormone (SH)-secreting cells, by means of histochemical and electron microscopic techniques. The newt testis is divided into two main parts of spermatogenetic stages. The anterior part is immature and consists of many seminal cysts containing spermatogonia and spermatocytes. The posterior part is mature and made up of many cysts containing mature spermatozoa and glandular tissue. Histochemical studies revealed Δ⁵-3β-hydroxysteroid dehydrogenase (Δ⁵-3β-HSD) activity in the pericystic cells of the mature part, weak in those before spermiation, then gradually becoming more intense in the pericystic cells just after spermiation. Activity was most pronounced in the outer layer of cells of the glandular tissue which originate from the pericystic cells. In contrast to this, the Δ⁵-3β-HSD reaction was entirely negative in the pericystic cells of the immature part, the Sertoli cells, and the inner cells of the glandular tissue originating from the Sertoli cells. Electron microscopic observation of the pericystic cells and glandular tissue gave further information on the histochemical features. The pericystic cells in the immature part are fibroblast-like cells. The cell organelles of these cells are poorly developed except for rough-surfaced endoplasmic reticulum and rod-shaped mitochondria. In the mature part, the fibroblast-like cells are transformed into cells characteristic of SH-secreting cells. In the cytoplasm, the amount of smooth-surfaced endoplasmic reticulum increases and typical globular mitochondria with tubular cristae take the place of the rod-shaped mitochondria. In addition, a number of lipid droplets and lipofuscin granules appear. Most of the fine structures of the outer layer of the cells of the glandular tissue were found to be similar in appearance to those of the SH-secreting cells in mammals and other vertebrates. These results strongly suggest that, as in the other urodeles, the SH-secreting cells in the testis of the Japanese red-bellied newt are the pericystic cells in the mature part and in the outer layer of cells of the glandular tissue.     

FORMATION OF GERM CELL-LIKE CELLS IN THE PERITONEAL EPITHELIUM OF THE TESTES OF ESTROGEN-TREATED ADULT MALES OF THE JAPANESE RED-BELLIED NEWT, CYNOPS PYRRHOGASTER PYRRHOGASTER

Columnar cells of the peritoneal epithelium in slender cords of the testes were examined in normal and estradiol benzoate-treated Japanese red-bellied newt, Cynops pyrrhogaster pyrrhogaster, by light and electron microscopy. In normal newts, the peritoneal epithelium covering the slender cord consists of columnar cells, which contain extraordinarily large, oval or spindle-shaped nuclei with conspicuous indentations. The nucleus contains chromatin granules and the cytoplasm is filled with numerous tonofilaments. The primordial germ cells are scattered throughout the slender cord, and each cell is surrounded by a few follicle cells. Between the germ cells and follicle cells there are microvilli-like processes. The nucleus of primordial germ cells is multilobate and has electron lucent areas, dispersed chromatin and several electron-dense nucleoli. In the lighter cytoplasm, the nuage material is found very near to nuclear pores, and is frequently seen among the mitochondria. The nucleolus-like body is not associated with other organelles. The primary spermatogonia have bilobate nuclei. It is remarkable that most of the cytoplasmic organelles are found in the deep nuclear indentations. The nuage material and nucleolus-like body are well developed in the cytoplasm. After treatment of newts with estradiol benzoate for one year, four types of cells can be distinguished in the peritoneal epithelium. One type is quite different from the columnar cells. These newly appeared cells are large and light in appearance. Their nucleus is highly lobate, and contains dispersed chromatin and several nucleoli with compact electron dense material in its periphery. The cells are characterized by the presence of nuage material and nucleolus-like bodies in the cytoplasm. There are microvilli-like processes between these cells and adjacent elongated cells. These ultrastructural characteristics of the light cells are very similar to those of primordial germ cells and/or primary spermatogonia in normal testes. These findings suggest that the light cells which appear in the peritoneal epithelium of the testes on administration of estrogen may be germ line cells.     

BEHAVIOR OF SECONDARY SPERMATOGONIA OF THE NEWT, CYNOPS PYRRHOGASTER, IN IN VITRO CULTURE

Several cell types migrated cut from small pieces of newt testes cultivated in vitro. Flat fibroblastic cells migrated out within a few days. Then, secondary spermatogonia, identified by the presence of germ cell-specific substances and by the shape and appearance of their nucleus and subcellular organelles, migrated out over the sheet of fibroblastic cells. Sertoli cells co-migrated with secondary spermatogonia, maintaining a similar cellular arrangement to that of testicular cells in vivo. Mitosis of secondary spermatogonia both in clusters and as single cells was frequent from the third day until about 2 weeks after inoculation. During mitosis, active and periodic rotation of chromosomes was observed. Identification of the cell types and studies on their behavior were performed by electron microscopy and phase contrast microscopy.     

CHANGES OF CHROMOSOMES DURING THE EARLY NEURAL DEVELOPMENT OF A JAPANESE NEWT, CYNOPS PYRRHOGASTER

The karyotype of Cynops pyrrhogaster was determined on the mitotic chromosomes in the presumptive neural area of an early gastrula. 24 chromosomes of a diploid set consisted of 8 metacentric and 4 submetacentric pairs. Individual chromosomes were identified on the basis of their morphology and characteristic C-binding patterns. Sex chromosomes were not identified. Total length of the haploid chromosome set in the presumptive neural area decreased remarkably from morulae to gastrulae, further continued to decrease up to neurulae and thereafter remained unchanged till tail-buds. Chromosome shortening occurring from morulae to gastrulae was accompanied with a prominent decrease in chromosome volume, keeping chromosome width constant. Shortening took place evenly along the longitudinal axis of a chromosome. When gastrulae and neurulae were compared concerning their positions of the appearance of the C-bands, the basic pattern remained unchanged. In certain chromosomes, the number of C-bands decreased as the result of their fusion, as gastrulae proceeded to neurulae.     

BRAIN INDUCTION IN VARIOUSLY AGED PRESUMPTIVE ECTODERMS BY HEAD ORGANIZER IN CYNOPS PYRRHOGASTER

Brain formation in variously aged presumptive ectoderms of Cynops pyrrhogaster under the influence of the head organizer was examined by the sandwich method. The head organizer was obtained from the middle portion of the archenteron roof at the slit-blastopore stage. The presumptive ectoderm was taken from 0- to 36-hr exogastrulae. Exogastrulae were prepared from the earliest gastrulae just before invagination (0-hr embryos). The presumptive neural plate overlying the archenteron roof used as organizer was cultivated in an envelope of belly ectoderm from an early neurula.
The following results were obtained: 1) Brain induction was almost entirely restricted to explants covered with 6-hr ectoderm and its frequency was low. 2) The presumptive neural plate above the head organizer was almost completely determined as neural tissues. 3) The head organizer showed a tendency to differentiate into more endodermal and less mesodermal tissues than those expected from its prospective fate.
Brain induction in normal development and the relationship between neural tissue formation in variously aged presumptive ectoderms and the time necessary for neural induction are discussed.     

'NUAGE'-CHROMOSOME ASSOCIATION IN METAPHASE SPERMATOGONIA OF THE NEWT, CYNOPUS PYRRHOGASTER

The 'nuage', an electron dense cytoplasmic component specific to the germ line cell, was found to be closely associated with a metaphase chromosome in the spermatogenic cell of the newt, Triturus (Cynops) pyrrhogaster. This finding suggests a possible relationship between both components at a certain stage of spermatogenesis.     

EFFECT OF AGING ON NEURAL COMPETENCE OF PRESUMPTIVE ECTODERM, AND EFFECT OF AGED ECTODERM ON DIFFERENTIATION OF THE ORGANIZER IN CYNOPS PYRRHOGASTER II. ROLE OF EXTREME POSTERIOR OF THE ARCHENTERIC ROOF IN THE SLIT-BLASTOPORE STAGE IN NORMAL DEVELOPMENT

The effect of aging on the neural competence of the presumptive ectoderm in gastrulae of Cynops pyrrhogaster and the effect of aged ectoderm on differentiation of the extreme posterior of the archenteric roof in the slit-blastopore stage were examined by a sandwich method in which this organizer was wrapped in the presumptive ectoderm taken from the 0- to 42-hr aged exogastrulae. Vital staining showed that this organizer becomes mainly tail notochord. Therefore it should be called tail or trunk-tail organizer. In 0- to 18-hr explants, typical trunk-tail structures were formed. With further aging of the presumptive ectoderm, a decrease of spinal cord and muscle with a concomitant increase of mesenchyme and mesothelium was observed. In 36- (corresponding to the slit-blastopore-initial neural stage) and 42-hr explants, neural competence had disappeared markedly. The notochord appeared in all explants, indicating this organizer is more firmly determined than the uninvaginated dorsal lip in small yolk-plug stage. Conclusively, this organizer does not play an important role in the induction of the neural plate, but induces the tail in normal development.     

A UNIQUE JUNCTIONAL STRUCTURE FOUND IN BLASTULA CELLS OF THE NEWT, TRITURUS PYRRHOGASTER

The outermost cell layer of the animal half of the newt blastula ( Triturus pyrrhogaster ) was examined to investigate intercellular junctions by transmission and scanning electron microscopy. A unique structure is observed at the terminal region of the intercellular junction. The structures are cytoplasmic ridges elevated from the cell surfaces, and their inner part is filled with spaces of various sizes. It is supposed that these ridges result from the encounter of cytoplasmic folds protruding from two neighboring cells.
Below the ridges, there is a short close junctional area which is followed by a long region of intercellular space intermittently bridged by cytoplasmic projections. Microvillus-like cytoplasmic processes on the apical cell surfaces, and microfilaments and microtubules in subsurface regions are observed in this material as in many other embryonic cells of amphibians.     

OCCURRENCE AND FINE STRUCTURE OF SUBCORTICAL CYTOPLASMIC ISLETS IN FERTILIZED EGGS OF CYNOPS PYRRHOGASTER

Fertilized and unfertilized eggs of Cynops pyrrhogaster were examined by light and electron microscopy. In fertilized eggs that have just been laid, there are numerous small cytoplasmic patches free of granules in the pigmented layer of the animal hemisphere. Many of these granule-free cytoplasmic islets gradually grow out subcortically from the pigmented layer and fuse to form a subcortical layer of yolk-free cytoplasm of varying thickness by the time of the first cleavage division. The cytoplasmic islets are present in 100% of the fertilized eggs, but not in unfertilized eggs. Electron microscopic observations showed that the cytoplasmic islets contain tubules and that development of a complex system of cortical tubules constitutes the basis of the early growth of the cytoplasmic islets. The cortical tubules are transient structures and are no longer observable a few hours after the eggs are laid. These phenomena are considered to be a response of the egg to the fertilization stimulus.     

东方蝾螈胚胎肌节发育过程中结蛋白的出现和分布

本文用间接免疫荧光方法观察了结蛋白在东方蝾螈胚胎发育过程中的生成时期和分布部位。结蛋白最早出现在分期25的蝾螈胚胎中,位于长柱形的肌细胞两端。随后肌细胞中结蛋白的含量随着发育逐渐增加,呈纤维状与细胞的纵长轴平行,但仍保持着两端含量高的特点。在分期41的胚胎中观察到结蛋白与Z线结合,晚于横纹的出现。根据分布特点讨论了结蛋白在肌细胞内的分布形式与脊索的关系,并对结蛋白从纤维状转变为和Z线相结合的过程以及结蛋白、肌小节的量的积累之关系作了讨论。     

INDUCING TIME FOR NEURAL TISSUES IN GASTRULA ECTODERM OF CYNOPS PYRRHOGASTER

The inducing times for spinal cord and deuterencephalon in Cynops gastrula were determined by the sandwich method. The extreme posterior of the archenteron roof at the slit-blastopore stage (tail organizer) was used as an inducer. First, the presumptive ectoderm of the earliest gastrula (0-hr stage) was put in contact with the organizer for 6 to 24 hr. Spinal cord and deuterencephalon were induced in almost all explants after 24 and 21 hr of contact, respectively, indicating that 24 hr is enough time for differentiations of both spinal cord and deuterencephalon. Next, presumptive ectoderm of 6- to 21-hr exogastrulae was put in contact with the organizer until the 24-hr stage. Results showed that the net inducing times for spinal cord and deuterencephalon were 18 and 15 hr, respectively, and that neural competence appeared in the presumptive ectoderm at the 6-hr stage (straight-blastopore stage).     

CORTICAL AND SUBCORTICAL CHANGES PRECEDING FURROW FORMATION IN THE CLEAVAGE OF NEWT EGGS

In the first cleavage of the egg of the newt, Cynops(Triturus) pyrrhogaster, some sort of preparation for the furrow formation in the cortical and subcortical cytoplasm precedes the advancing tip of the cleavage furrow. This is shown by the following facts: (1) Incisions made close to the tip of the cleavage furrow do not stop the progress of furrowing, allowing the furrow to cross the incisions and appear on the farther sides, while incisions made far enough from the furrow tip always prevent the further travelling of the furrow, (2) Displacement of the subcortical cytoplasm ahead of the furrow by rubbing with a hair loop makes the furrow bend corresponding to the width of the rubbed area, and (3) Transplantation of the subcortical material of the furrow tip to lateral parts in the same egg causes a depression in the overlying cortex at the transplanted position. The linear extension of the prepared area for the furrow formation is the longest in the animal hemisphere and it decreases in gradient towards the vegetal pole.     

NPY样免疫反应在蝾螈发育过程中神经系统的定位研究

用免疫组化方法研究在蝾螈胚胎发育中神经肽Y(NPY)在神经系统中的定位.结果表明NPY最早分布在周边神经系统(尾芽晚期),之后才在中枢神经系统出现(幼虫期),而且是随着胶质细胞的出现而出现的.我们认为NPY是随着神经嵴细胞的迁移而进入周边神经系统,再分布到中枢神经系统的.     

侧柏小孢子的发生和雄配子体的形成

侧柏[Platycladusorientalis（L．）Franco]初生造孢细胞在8月下旬（1992年）形成,11月上旬形成小孢子母细胞,1993年2月中旬形成四分体,2月下旬小孢子从四分体内释放出来,3月中旬形成成熟花粉粒并开始传粉,4月上旬花粉粒在珠心上萌发,5月上旬生殖细胞分裂,6月上旬精原细胞分裂。小孢子母细胞在休眠以前开始减数分裂,解除休眠以后形成成熟的花粒粒。减数分裂从11月上旬开始至次年2月17日结束。小孢子母细胞减数分裂存在扩散双线期。小孢子母细胞以双线期渡过休眠。精原细胞接近颈卵器时开始分裂,形成两个大小相同的精细胞。精细胞独立存在的时间很短。精细胞的细胞质分为三个区域。小孢子母细胞在发育过程中,发现有部分小孢子母细胞退化,在小孢子囊内形成一大的空腔的现象。     

啤酒酵母原生质体的制备、融合及再生

本文以16号(his~-)啤酒酵母和STA(ade~-)啤酒酵母作出发菌株进行原生质体融合,并探讨了菌龄、酶解时间,菌体分散程度和培养基种类等对原生质体融合的影响。     

日本沼虾输精管的结构及其在精荚形成中作用的研究

应用光镜和透射电镜技术研究了日本沼虾输精管的结构及其在精荚形成中的作用。结果表明,日本沼虾输精管从形态结构上可分为近端输精管、卷曲输精管、远端输精管和膨大的远端输精管四部分。各部分的管壁皆由分泌上皮、基膜、肌肉层和结缔组织构成,其中分泌上皮包括高度明显不同的低柱状上皮和高拄状上皮两部分。输精管各部分管腔内含有处于不同形成阶段的精荚。进入近端输精管内的精子被支撑在一种嗜酸性基质中。近端输精管的分泌物主要帮助形成精子团,同时形成精荚壁的极小部分。卷曲和远端输精管分泌形成精荚壁的绝大部分,其分泌物由细胞顶端通过外排作用和顶泌机制分泌产生。膨大的远端输精管具有贮存精荚的作用,其分泌上皮也通过外排作用和顶泌机制产生分泌物包裹在已基本形成的精荚外侧,管壁肌肉层在雌雄交配时将管腔内的精荚切割成适宜长度并排出体外。       

有尾两栖类细胞连接和神经管形成

在神经管形成的过程中,从原肠晚期到神经管即将关闭,我们观察了预定神经上皮和神经上皮细胞的细胞连接。我们的结果和过去的报道不同,我们看到紧密连接和间隙连接在所观察的各个时期一直存在,而且它们是处于持续的形态变化之中,就此论证了紧密连接在神经管形态建成中的作用,并讨论了紧密连接和间隙连接的相互关系。     

试论中国第三果树带的形成与发展

本文提出了果树带的新概念,即根据果树栽培的高效优质及其自然分布情况,提出在长江以南（北纬１８─３２°）柑桔为主的第一果树带和长江以北（北纬３２─４３°）苹果为主的第二果树带之间,即北纬２５─３５°建立和发展以猕猴桃为主的第三果树带的新论点。同时,因海拔高度的变化以及在两带的边缘地域,表现出部分过度交互型特征。本文还进一步阐述了以猕猴桃为代表的第三果树带形成和发展的可能与条件,指出提高果树种植效益与不同果树相应气候带的关系,分析说明了中国第三果树带的形成与发展的重要意义。此外,还对第三果树带的果树组成、发展方向和途径等关键性问题进行了讨论,为中国第三果树带的形成与发展提供了理论依据。     

轮虫休眠卵形成和萌发的生态机理研究进展

休眠卵的形成是单巢目轮虫生殖行为中最显着的特征之一。作为一系列有性生殖过程的产物,休眠卵具有抵抗外界不良环境、易于扩散和通过重组产生遗传多样性等特点。轮虫休眠卯是轮虫集约化培养的重要种源,通过轮虫快速的孤雌生殖在短时间内大量繁殖,使得轮虫作为鲜活饵料为水产经济动物的苗种生产起到了极其重要的作用。但是在生产实践中,利用孤雌生殖批量培养轮虫常会因难以预测的种群突然崩溃而造成水产经济动物苗种生产的重大损失。