Theory and practice of stereological techniques applied to the estimation of cell number and nuclear volume in the testis.
ABSTRACT The historical background to contemporary approaches to the estimation of cell/nuclear number and volume in the testes is reviewed. The limitations of older geometric model-based approaches to the estimation of cell/nuclear number are discussed, and the need for absolute estimates of cell number rather than ratio estimates is examined. The physical and optical disector approaches to the direct estimation of numerical density and, hence, absolute cell number are presented together with data illustrating their operational efficiency in the testis. New approaches to the direct estimation of nuclear/cell volume, using the point-sampled intercept family of methods, are presented and illustrated, using the example of the Sertoli cell nucleus. The use of both classical transverse and the newer vertical section approaches is explored. Estimation of Sertoli cell/nuclear volume in the volume (point-sampled intercept procedure) and number (nucleator and rotator methods) distributions on both conventional transverse and vertical sections is discussed. The use of transverse sections of the testis is shown to produce a consistent bias in the estimation of Sertoli cell nuclear volume in 120-day-old animals, with all the estimators. Comparison of the Sertoli cell nuclear volume (measured on vertical sections) in the volume and number-weighted distribution suggests a coefficient of variation of volume in the number distribution of 0.4-0.5, suggesting either a random or stage-dependent variation in Sertoli cell nuclear size which requires further exploration.
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ABSTRACT: The Sertoli cells are critical regulators of testis differentiation and development. In the adult, however, their known function is restricted largely to maintenance of spermatogenesis. To determine whether the Sertoli cells regulate other aspects of adult testis biology we have used a novel transgenic mouse model in which Amh-Cre induces expression of the receptor for Diphtheria toxin (iDTR) specifically within Sertoli cells. This causes controlled, cell-specific and acute ablation of the Sertoli cell population in the adult animal following Diphtheria toxin injection. Results show that Sertoli cell ablation leads to rapid loss of all germ cell populations. In addition, adult Leydig cell numbers decline by 75% with the remaining cells concentrated around the rete and in the sub-capsular region. In the absence of Sertoli cells, peritubular myoid cell activity is reduced but the cells retain an ability to exclude immune cells from the seminiferous tubules. These data demonstrate that, in addition to support of spermatogenesis, Sertoli cells are required in the adult testis both for retention of the normal adult Leydig cell population and for support of normal peritubular myoid cell function. This has implications for our understanding of male reproductive disorders and wider androgen-related conditions affecting male health.PLoS ONE 01/2014; 9(8):e105687. · 3.53 Impact Factor
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ABSTRACT: The AS/AGU rat is a spontaneous recessive mutation derived from a closed colony of Albino Swiss (AS) rats. This mutation generates a stop codon within the coding region of the PKC-gamma gene. The rats exhibit locomotor dysfunc-tion, which is progressive with age. The nigrostriatal system has already been shown to be affected by this mutation that a) there is marked (80-90%) reduction in extracellular dopamine in the dorsal caudate-putamen as revealed by in vivo microdialysis and HPLC-ECD and b) there is a loss of tyrosine hydroxylase immunoreactive cells in the substantia nigra pars compacta Because PKC-gamma may be involved in the packaging or release of vesicles, and because other aminergic neurons may also be involved in basal ganglia disorders, we investigated the raphe-striatal serotonergic sys-tem in these mutants and the parent AS strain. Animals aged 12-15 months (6 pairs) were killed by administration of sodium pentobarbitone and were perfused with Ringer's solution + Lignocaine followed by 4% paraformaldehyde. 300m sections were cut, rostral to caudal, in a one in five series. The collected sections were processed for immu-nostaining using the Avidin-Biotin-Peroxidase Complex (ABC) technique. All sections of dorsal raphe and median raphe nuclei were counted using the light microscope and also by unbiased Sterological methods using "Stereologer". In each case, AS/AGU rats possessed an average of 20-25% fewer cells in the dorsal raphe than AS control rats (p<0.05); cell count in the median raphe were unaffected.
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ABSTRACT: To investigate the effects of a low bisphenol A (BPA) concentration on male reproduction, adult rats were administered a concentration of BPA that was less than the no observable adverse effect level (0.0005–5 mg/kg/bw) for 8 weeks. General toxicity, reproductive hormones, and spermatogenesis were then determined. The expression of genes related to hormone synthesis and spermatogenesis was also analyzed. These BPA concentrations generated no general toxicity and no significant changes on serum hormones. However, the testicular testosterone, hormone synthesis-related genes StAR and Cyp450scc increased, whereas 3β-HSD, 17β-HSD, and Cyp450arom decreased. Additionally, BPA significantly decreased the epithelial height and round spermatids in seminiferous tubules, sperm count, androgen receptor expression, and the expression of the spermatogenesis-related genes outer dense fiber protein 1 (ODF1) and transition protein 1. Our results indicate that a low BPA concentration can induce spermatogenesis disorders mainly through decreasing androgen receptor expression. The present results may bring attention to the risk of environmental BPA exposure.Toxicology Letters 05/2013; 219(2):116–124. · 3.36 Impact Factor