Pyrimethamine(PYR) produces reversible infertility in mice. The antifertility effect of PYR is due to its antifolate action. PYR causes male infertility in Balb C mice in a dose dependent manner. The present study was performed to determine DNA synthesis of germ cell in mice treated with PYR. For 6-8 weeks adult Balb C mice were administered a PYR dose of 100mg- 200mg/ kg/day suspended in vegetable oil.10 control mice received only vegetable oil without PYR. After 7 weeks administration 28.6% of mice(4/14) administered a PYR dosage of 100mg/kg were infertility, whereas all of those(4/4), 200mg/kg/day were infertile. Morphologic changes of the testis induced by orally administered PYR were spermatogenic arrest and depopulation of germ cell. The incorporation of thymidine into DNA of germ cells was studied by using autoradiography after intraperitoneal injection of 15 Ci of methyl-3H-thymidine. Synthesis of DNA takes place in interphase and early prophase of mitotic phase of germ cell. The synthesis of DNA in spermatogonia reduced in all stage of spermatogenesis in mice treated with PYR due to spermatogenic arrest, compared with that of control mice. These results suggest that reduced synthesis of DNA in spermatogonia by PYR administration caused hypospermatogenesis. PYR represents further approach toward development of male contraceptive.
Adult ; Animals ; Autoradiography ; DNA* ; Germ Cells* ; Humans ; Infertility ; Infertility, Male ; Injections, Intraperitoneal ; Interphase ; Male ; Mice* ; Mice, Inbred BALB C ; Oligospermia ; Prophase ; Pyrimethamine* ; Spermatogenesis ; Spermatogonia ; Testis* ; Thymidine ; Vegetables

Adult ; Azoospermia/pathology* ; DNA Breaks, Double-Stranded ; DNA Repair ; Humans ; Infertility, Male/pathology* ; Male ; Prophase ; Prostate/pathology* ; Recombination, Genetic ; Synapses/pathology* ; Testis/pathology*

Chfr, a mitotic stress checkpoint gene, regulates a prophase delay in cells exposed to agents that disrupt microtubules, such as nocodazole and taxol. Chfr expression was ubiquitious in normal human tissues. It is very high conserved between human and mice. Preliminary sutdies indicated that Chfr expression was cell cycle regulated and it dependent on its ubiqitin ligase activity. The direct target of the Chfr pathway was Polo-like kinase 1 (Plk1). Ubiquitination of Plk1 by Chfr delayed the activation of the Cdc25C phosphatase and the inactivation of the Weel kinase, leading to a delay in Cdc 2 activation. The chfr gene was inactivated owing to lack of expression or by mutation in some human cancer cell lines examined. Normal primary cells and tumour cell lines that express wild-type chfr exhibited delayed entry into metaphase when centrosome separation was inhibited by mitotic stress. In contrast, the tumour cell lines that had lost chfr function entered metaphase without delay. Ecotopic expression of wild-type chfr restored the cell cycle delay and increased the ability of the cells to survive mitotic stress. Thus, chfr defines a checkpoint that delays entry into metaphase in response to mitotic stress. The progress of research on structure of Chfr gene and effects of Chfr protein was reviewed.
Cell Cycle ; genetics ; physiology ; Cell Cycle Proteins ; genetics ; metabolism ; physiology ; Humans ; Metaphase ; genetics ; physiology ; Mitosis ; genetics ; physiology ; Neoplasm Proteins ; genetics ; physiology ; Neoplasms ; genetics ; metabolism ; pathology ; Poly-ADP-Ribose Binding Proteins ; Prophase ; genetics ; physiology ; Protein-Serine-Threonine Kinases ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Ubiquitin-Protein Ligases

The synaptonemal complex (SC) is a meiosis-specific proteinaceous macromolecular structure that assembles between paired homologous chromosomes during meiosis in various eukaryotes. The SC has a highly conserved ultrastructure and plays critical roles in controlling multiple steps in meiotic recombination and crossover formation, ensuring accurate meiotic chromosome segregation. Recent studies in different organisms, facilitated by advances in super-resolution microscopy, have provided insights into the macromolecular structure of the SC, including the internal organization of the meiotic chromosome axis and SC central region, the regulatory pathways that control SC assembly and dynamics, and the biological functions exerted by the SC and its substructures. This review summarizes recent discoveries about how the SC is organized and regulated that help to explain the biological functions associated with this meiosis-specific structure.
Animals ; Chromosome Segregation ; Meiosis/physiology* ; Synaptonemal Complex/physiology*

Animals ; Centromere ; metabolism ; Chromosomal Proteins, Non-Histone ; metabolism ; Mice ; Models, Molecular ; Synaptonemal Complex ; metabolism

Successful male germ cell transplantation requires depletion of the host germ cells to allow efficient colonization of the donor spermatogonial stem cells. Although a sterilizing drug, busulfan, is commonly used for the preparation of recipient models before transplantation, the optimal dose of this drug has not yet been defined in dogs. In this study, 1-year-old mongrel dogs were intravenously injected with three different concentrations of busulfan (10, 15, or 17.5 mg/kg). Four weeks after busulfan treatment, no fully matured spermatozoa were detected in any of the busulfan-treated groups. However, small numbers of PGP9.5-positive spermatogonia were detected in all treatment groups, although no synaptonemal complex protein-3-positive spermatocytes were detected. Of note, acrosin-positive spermatids were not detected in the dogs treated with 15 or 17.5 mg/kg busulfan, but were detected in the other group. Eight weeks after busulfan treatment, the dogs treated with 10 mg/kg busulfan fully recovered, but those in the other groups did not. PGP9.5-positive spermatogonia were detected in the 10 mg/kg group, and at a similar level as in the control group, but these cells were rarely detected in the 15 and 17.5 mg/kg groups. These results suggest that a dose of 15-17.5 mg/kg is optimal for ablative treatment with busulfan to prepare the recipient dogs for male germ cell transplantation. At least eight weeks should be allowed for recovery. The results of this study might facilitate the production of recipient dogs for male germ cell transplantation and can also contribute to studies on chemotherapy.
Animals ; Busulfan* ; Colon ; Dogs* ; Drug Therapy ; Germ Cells ; Humans ; Male ; Spermatids ; Spermatocytes ; Spermatogonia ; Spermatozoa ; Stem Cell Transplantation* ; Stem Cells* ; Synaptonemal Complex ; Testis* ; Tissue Donors

OBJECTIVETo analyze defective homologous chromosomal recombination in Han Chinese azoospermic patients.

METHODSTesticular biopsy samples from 7 healthy controls and 7 Han Chinese azoospermic patients including 2 obstructive azoospermia (OA group) and 5 non-obstructive azoospermia (NOA group) were analyzed. Immunofluorescence staining was performed to categorize early stage cells at meiosis prophase and to analyze chromosome pairing and recombination of pachytene spermatocyte. Newly developed meiotic proteins antibodies (anti-SCP3, anti-synaptonemal complex proteins 3, anti-MLH1, anti-Mut-L Homolog 1, anti-CREST, chromosome centromere antibody) were used to identify synaptonemal complex (anti-SCP3), recombination sites (anti-MLH1) and centromere (anti-CREST), respectively. Staging of spermatocyte was determined according to SCP3 formation progression. Qualitative data were compared by a Chi-square test, and ANOVA was used to analyze quantitative data.

RESULTSRespectively, 2346 and 2932 spermatocytes were categorized in the controls and azoospermic patients. The proportions of zygotene cells in both OA group and NOA group were significantly higher than that of the control group. Investigation of 1967 pachytene cells from the controls and 354 pachytene cells from azoospermic patients indicated that the mean MLH1 foci per pachytene cell of NOA group was statistically lower than that of the controls. Compared with the controls, incomplete synaptonemal complexes cells (containing gap and/or split) were significantly increased in the NOA group.

CONCLUSIONDelayed meiosis prophase is relatively common in azoospermic patients, and changes in quantity and distribution of recombination foci may be the cause for spermatogenesis arrest in Han Chinese population.

Adult ; Asian Continental Ancestry Group ; Azoospermia ; genetics ; metabolism ; pathology ; Humans ; Male ; Meiosis ; genetics ; Middle Aged ; Recombination, Genetic ; Spermatocytes ; metabolism ; Synaptonemal Complex ; genetics ; Young Adult

Introduction of patch-clamp techniques allowed an increase in resolution of membrane current recordings. However, the technique was limited by apparent need for direct contact of pipette with cell membrane. Thus, this technique was restricted to isolated or cultured cell preparation. Although much has been achieved with such preparations, the studies of synapsis between cultured cells are undefined. Many of these problems were overcome by application of patch-clamp techniques to brain-slices. The use of high-resolution optics allowed visualization of cells to be recorded. It was possible to remove tissue covering cells and record currents in synaptically connected neurons. The brain-slice technique has greatly facilitated the investigation of electrical properties of neurons and the analysis of synaptic transmission between neurons. "Blow and seal"technique, when combined with infrared differential interference contrast video microscopy, permits recording of membrane potential and currents, not only from large cell body of neurons, but also from small processes. The technique offers many advantages, such as the case with which patch-pipette recordings can be made, the possibility of identifying cell type prior to recording and finally, the ability to visualize and record electrical activity from different compartments or from more than one site in the same neuron.
Anesthetics, General ; Cell Membrane ; Cells, Cultured ; Chromosome Pairing ; Membrane Potentials ; Membranes ; Microscopy, Video ; Neurons ; Patch-Clamp Techniques ; Synaptic Transmission

To investigate the distribution, ultrastructure and synapsis of serotoninergic cells and CGRP nerve fibers in mammalian taste buds, immunohistochemistry and electronmicroscopy were applied to mice vallate papillae. In normal mice, 1~2 serotonin immunoreactive cells were present in each taste bud section. After preloading 5-HTP, 3~6 cells showed strong immunoreactivity for serotonin. These cells were elongated, and their cytoplasm extended from the taste pore to the base of the taste bud. CGRP nerve fibers formed thick subgemmal nerve plexus under the basal lamina, and branched varicose perigemmal and intragemmal nerve fibers. Under the electron-microscope, three types of taste cells; dark cell, light cell and basal cells, were identified by their shape, location and electrical densities. Immuno-electronmicroscopy revealed that serotoninergic cells were dark cells. CGRP nerve fibers were located in and around taste buds, but the synaptic contacts with taste cells was not found. These findings indicate that mice taste cells are consisted of dark cell, light cell and basal cells, and dark cells contain serotonin. And, CGRP nerve fibers in taste buds may function as general sensory fibers.
5-Hydroxytryptophan ; Animals ; Basement Membrane ; Calcitonin Gene-Related Peptide* ; Calcitonin* ; Chromosome Pairing ; Cytoplasm ; Immunohistochemistry ; Mice* ; Nerve Fibers* ; Serotonin ; Taste Buds*

A rare case of intraventricular central neurocytoma in 17-year-old male is reported. The patient had diffuse headache and diplopia. Radiologic findings displayed obstructive hydrocephalus and a large, well-demarcated intraventricular mass lesion obstructing the foramen of Monroe. The tumor arouse from the splenium of corpus callosum. It was removed successfully using two different approaches after extraventricular drainage of the cerebrospinal fluid(CSF). Histologically, the tumor showed pathological features as that of oligodengroglioma on the light microscope. In immunohistochemical examination, glial fibrillary acidic protein(GFAP) was negative and synaptophysin, positive. Numerous neurosecretory granules were found and no typical synapsis was noticed on the electron microscope. No shunt operation was needed. Postoperative radiotherapy or chemotherapy was not performed and no tumor recurrence was detected during the one year follow-up period. We present the case together with a review of the literatures.
Adolescent ; Cerebral Ventricles ; Chromosome Pairing ; Corpus Callosum ; Diplopia ; Drainage ; Drug Therapy ; Follow-Up Studies ; Headache ; Humans ; Hydrocephalus ; Male ; Neurocytoma* ; Radiotherapy ; Recurrence ; Synaptophysin