Seminiferous tubule transfection in vitro to define post-meiotic gene regulation

Background  

Post-meiotically expressed genes in the testis are essential for the proper progression of spermatogenesis, and yet, aside from the construction of individual transgenic mice using specific promoters to drive reporter plasmids, there are only very limited possibilities for relevant and quantitative analysis of gene promoters. This is due to the special nature of post-meiotic haploid cells, which to date are not represented in any appropriate cell-lines. This article reports the development of novel methodology using isolated and cultured rat seminiferous tubules in a multiwell format, into which promoter-reporter constructs can be introduced by a combination of microinjection and electroporation.     

The incidence of Weeksella- and Bergeyella-like bacteria in the food environment

Eighty-five catalase- and oxidase-positive Gram-negative rods and cocci susceptible to penicillin G were isolated from a variety of food sources. The phenotypic relationships of these isolates with reference cultures of Bergeyella -like, Chryseobacterium, Empedobacter, Myroides , Moraxella , Sphingobacterium and Weeksella -like strains were examined by numerical taxonomy. Seventy-three isolates were recovered in five groups; 80% of the isolates clustered in groups 1, 2 and 3 and produced indole, bearing a strong resemblance to Weeksella and Bergeyella . They could not, however, be regarded as belonging to the known species of W. virosa and B. zoohelcum . It is suggested that three species may be necessary to accommodate the environmental Weeksella - or Bergeyella -like bacteria. The isolates in groups 4 and 5 had white colonies and were unable to produce indole, in this way resembling the Moraxella genus.     

Correlation between the parameters of contingent negative variation and characteristics of variational pulsometry in Parkinsonian patients

In patients suffering from Parkinson's disease (PD), we analyzed correlations between the parameters of contingent negative variation (CNV) and data of variational pulsometry (according to the measurements of R-R ECG intervals). Studies were carried out on 35 patients (group PD), 49 to 74 years old, with the stage of disease of 1.5 to 3.0 according to the Hoehn-Yahr international classification. In the course of CNV recording (i.e., in the state of a certain functional loading), we observed significant negative correlations between the integral magnitude (area) of this potential and indices of variational pulsometry (RMSSD, SDNN, C. var, and HF) that characterize the intensity of parasympathetic (respiratory) influences on the cardiovascular system. In the control group, such correlations were absent. We found significant correlations between the autonomic balance, CNV magnitude, and stage of PD reflecting the level of generalization of the pathological process. In the subgroup of patients with the PD stage 1.5 to 2.0, significant changes in the mean values of indices of parasympathetic influences during recording of the CNV were not observed, while in another subgroup (the PD stage 2.5 to 3.0), these values increased significantly (P < 0.05 and P < 0.01). If the estimates of the PD stage were low, the CNV area demonstrated greater values (P < 0.01). The disturbance of coordination of muscle-to-muscle interactions in the PD group is, probably, an important factor responsible for parasympathetic dysregulation and suppression of the CNV generation. We found positive correlation between the intensity of parasympathetic influences in the course of CNV recording and the level of postural disorders (r = 0.37, P < 0.05). On the contrary, the CNV magnitude demonstrated a negative correlation with the intensity of these disorders (r = −0.36, P < 0.05), as well as with the level of postural instability (r = −0.55, P < 0.001). We hypothesize that alterations of the autonomic balance and the activity of those cerebral structures, which are responsible for the motor readiness, result, to a significant extent, from weakening of the activity of the noradrenergic system due to degenerative processes developing in cells of the locus coeruleus. The impairment of the latter structure, together with degeneration of neurons of the substantia nigra and a decrease in the level of nigro-striatal dopamine, underlies the pathomorphological pattern of PD. Neirofiziologiya/Neurophysiology, Vol. 40, No. 3, pp. 242–253, May–June, 2008.     

Immunofluorescent analysis of meiotic recombination in the domestic cat

The paper presents the analysis of the frequency, density, and distribution of recombination sites in the male meiosis of the domestic cat (Felis silvestris catus). The study was carried out using immunofluorescent staining of synaptonemal complex (SC) proteins, centromeric proteins and mismatch repair protein MLH1, a reliable marker of crossingover sites. We mapped 2633 sites of crossing over in 1098 individual autosomes. Based on these data, we estimated the total length of the genetic map of the domestic cat to be 2176 centimorgans. Positive correlation between the length of SC and the number of recombination sites common for mammalians was also found in the domestic cat. It was shown that this species was characterized by the highest density of recombination and the lowest interference in mammals.     

Differentiation of the myoepithelial cells of the rat submandibular gland in vivo and in vitro: an ultrastructural study

Cytodifferentiation of the myoepithelial cells (MEC) of the rat submandibular gland (SMG) was observed by studying the prenatal and postnatal development of the gland in vivo and in vitro by light and electron microscopy. The anlage of the SMG first appeared on the fourteenth day of gestation and, from its earliest inception, was surrounded by an intact basal lamina. Presumptive myoepithelial cells were first seen at 18 days of gestation coinciding with the onset of secretion in the rudiment. These cells were flattened, peripherally located and subjacent to the epithelial basal lamina. Initial deposition of cytofilaments in the MEC's was observed during the first three days following birth and fully matured cells were seen as early as one week after birth. Presumptive and immature MEC's were observed undergoing mitosis, but once cytofilament deposition had begun in the cells they did not divide. Myoepithelium developed in relation to embryonic secretory structures and were only observed surounding acini and intercalated ducts in the adult gland. New myoepithelial cells were formed as long as new acinar-intercalated duct units were formed. Myoepithelial cells did not produce secretory type granules at any time during their development or in their mature state. Development of the MEC's in vitro paralleled that in vivo and supported the above observations.     

An approach to the focal presentation of chemical stimuli

An olfactometer is described that presents temporally-discretepulses of stimuli to individual chemosensory structures. Thedevice is based on standard pressure injection techniques inwhich pulses of compressed air eject nanoliter volumes of upto six stimulants from small diameter glass micropipettes. Thedevice should be readily adaptable to different chemoreceptorpreparations.