Increase in c-Fos and Arc protein in retrosplenial cortex after memory-improving lateral hypothalamic electrical stimulation treatment.

Post-training Intracranial self-stimulation (ICSS) of the lateral hypothalamus (LH), a kind of rewarding deep-brain stimulation, potentiates learning and memory and increases c-Fos protein expression in specific memory-related brain regions. In a previous study, Aldavert-Vera et al. (2013) reported that post-acquisition LH-ICSS improved 48 h retention of a delay two-way active avoidance conditioning (TWAA) and induced c-Fos expression increase in CA3 at 90 min after administration. Nevertheless, this c-Fos induction was only observed after the acquisition session and not after the retention test at 48 h, when the ICSS improving effect was observed on memory. This current study aims to examine the hypothesis that post-training ICSS treatment may stimulate c-Fos expression at the time of the TWAA retention test in retrosplenial cortex (RSC), a hippocampus-related brain region more closely related with long-lasting memory storage. Effects of ICSS on Arc protein, a marker of memory-associated synaptic plasticity, were also measured by immunohistochemistry in granular and agranular RSC. The most innovative results are that the ICSS treatment potentiates the c-Fos induction across TWAA conditions (no conditioning, acquisition and retention), specifically in layer V of the granular RSC, along with increases of Arc protein levels in the granular but not in agranular areas of RSC ipsilaterally few hours after ICSS. This leads us to suggest that plasticity-related protein activation in the granular RSC could be involved in the positive modulatory effects of ICSS on TWAA memory consolidation, opening a new approach for future research in ICSS memory facilitation.

Intracranial self-stimulation facilitates active-avoidance retention and induces expression of c-Fos and Nurr1 in rat brain memory systems.

Intracranial self-stimulation (ICSS), a special form of deep brain stimulation in which subjects self-administered electrical stimulation in brain reward areas as the lateral hypothalamus, facilitates learning and memory in a wide variety of tasks. Assuming that ICSS improves learning and memory increasing the activation of memory-related brain areas, the present work examined whether rats receiving an ICSS treatment immediately after the acquisition session of a two-way active avoidance conditioning (TWAA) show both an improved retention and a pattern of increased c-Fos and Nurr1 protein expression in the amygdala, hippocampus, dorsal striatum and/or lateral hypothalamus. The response of both activity-induced IEGs to ICSS was examined not only as markers of neural activation, but because of their reported role in the neural plasticity occurring during learning and memory formation. Results showed that the TWAA conditioning alone increased the expression of the two analysed IEGs in several hippocampal areas, and TWAA retention increased Nurr1 expression in amygdala. ICSS treatment increased the number of c-Fos and Nurr1 positive cells in almost all the brain regions studied when it was measured 70min, but not 48h, after the stimulation. Post-training ICSS treatment, as expected, facilitated the 48h retention of the conditioning. It is noteworthy that in CA3 conditioning and ICSS separately increased c-Fos expression, but this increasing was greater when both, conditioning and ICSS, were combined. Present results suggest that rapid and transient increased expression of these two synaptic plasticity and memory related IEGs in some hippocampal areas, such as CA3, could mediate the facilitative effects of ICSS on learning and memory consolidation.

High-frequency stimulation of the ventrolateral thalamus regulates gene expression in hippocampus, motor cortex and caudate-putamen.

High-frequency stimulation (HFS) of the ventrolateral (VL) thalamus is effective in treating the resting tremor of Parkinson's disease (PD). PD is a movement disorder that involves neurodegeneration, predominantly of the substantia nigra, but also in other brain areas, such as the motor cortex and hippocampus. The mechanisms of action of HFS on remote brain areas at the molecular level are largely unknown. Here, we investigated gene expression profiles using oligonucleotide microarrays and quantitative real-time PCR in rat hippocampi. We showed that chronic (14days) HFS modulates the expression of 176 hippocampal genes. Our results showed that genes involved in proliferation and neurogenesis-related biological functions were specifically regulated by HFS, including nestin (Nes) and doublecortin (Dcx), which are expressed in neural progenitor cells and immature neurons, respectively, as well as genes encoding proteins that may support neural differentiation or migration, such as Timp1, Ccl2, S100a4 and Angpt2. Next, we used quantitative real-time PCR (RT-PCR) to profile these six genes in the motor cortex and the caudate-putamen, which included the subventricular zone (CPu-SVZ). Interestingly, HFS increased Dcx expression in the motor cortex whereas Nes was upregulated in the CPu-SVZ but not in the motor cortex. In the CPu-SVZ Timp1 and Ccl2 were highly upregulated by HFS. In conclusion, our findings suggest that HFS may enhance neuroplasticity at the molecular level in several remote brain areas such as the CPu-SVZ, motor cortex and hippocampus.

Transgene expression levels determine the immunogenicity of transduced hematopoietic grafts in partially myeloablated mice.

We investigated whether transgene expression levels influence the immunogenicity of transduced hematopoietic grafts upon transplantation into partially myeloablated mice. To this aim, bone marrow cells (BMCs) transduced with retroviral vectors driving green fluorescent protein (GFP) expression either at high (high-EGFP) or low levels (low-EGFP) were transplanted into congenic recipients conditioned with sublethal doses of total body irradiation (TBI) or busulfan. Virtually all recipients showed evidence of donor engraftment 4 weeks after transplantation. However, as opposed to recipients receiving low-EGFP transduced grafts, the risk of rejecting the EGFP(+) cells by 30 days after transplantation was significantly higher in mice conditioned with busulfan and receiving high-EGFP transduced grafts. Anti-EGFP cellular immune responses were demonstrated in high-EGFP-treated mice conditioned with busulfan by interferon-gamma (IFN-gamma), enzyme-linked immunospot assay (ELISPOT), and cytotoxic T lymphocyte (CTL) assays, in contrast to that observed in mice transplanted with low-EGFP BMC. These results show for the first time that transgene expression levels can be critical for the immunogenicity of gene-modified hematopoietic grafts, especially in immunocompetent or in partially immunosuppressed recipients. These results have profound implications in vector choice and in the design of gene therapy (GT) protocols.

Effects of a high semen-collection frequency on the quality of sperm from ejaculates and from six epididymal regions in boars.

This study examines the effect of semen-collection rhythm on the sperm maturation process in boar epididymis. Three post-pubertal boars were submitted to a high semen-collection frequency (stressed boars) during 4 days, and three males were kept as a control group (control boars). Semen samples coming from six epididymal regions and from the ejaculate of each male were evaluated for sperm concentration, sperm vitality, sperm motility and sperm morphology. In each epididymal region, either fluid resorption or fluid secretion was determined from the variation in sperm concentration. The pattern of fluid resorption-secretion along the epididymal duct differed significantly between the stressed and control boars. A high semen-collection frequency also affected the development of sperm motility and the sperm cytoplasmic droplet displacement along the epididymal duct. The incidence of some sperm abnormalities was also found to be higher in some epididymal regions and ejaculates of stressed boars. From the results of this study, it can be concluded that a high semen-collection frequency brings about an altered resorption and secretion pattern of the epididymal fluid, which results in defective sperm maturation and abnormal development of sperm motility.

In vitro culture of epithelial cells from the caput, corpus, and cauda epididymis of Sus domesticus.

This work describes a protocol to culture epididymal epithelial cells from the caput, corpus, and cauda regions of Sus domesticus. Epididymal epithelial fragments were obtained by dissection and enzymatic digestion with collagenase. About 30 epididymal fragments from each epididymal region were cultured in 24-well culture plates with supplemented RPMI-1640 medium at 37 degrees C, 5% CO2 in air, and 100% humidity. A confluent monolayer of polygonal and tightly packed epithelioid cells from the three epididymal regions was obtained after 12-16 days in culture and maintained in vitro for more than 60 days. The proportion of epididymal epithelial cells in these cultures was assessed by immunofluorescent staining for cytokeratins. Throughout the 2 months of culture, about 80% of the cells were cytokeratin-positive. Electron microscopy observations indicated that cultured cells from caput, corpus, and cauda epididymal regions were tightly adhered to each other by junctional complexes and that stereocilia were present in their apical membranes. Moreover, the presence of an extensive rough endoplasmic reticulum, Golgi apparatus and numerous vesicles in the cytoplasm suggested that cultured cells maintained secretory and absorptive activities. These results show that the epididymal epithelial cells in culture from S. domesticus retain some fundamental features that characterize the epididymal epithelium in the intact organ. This system might be a valuable tool for studying the mechanism of sperm maturation in vitro, including epididymal cell secretions and the analysis of regional differences.