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1.
Glucose-stimulated calcium dynamics in Islets of Langerhans in acute mouse pancreas tissue slices
Andraž Stožer, Jurij Dolenšek, Marjan Rupnik, 2013, izvirni znanstveni članek

Opis: In endocrine cells within islets of Langerhans calcium ions couple cell stimulation to hormone secretion. Since the advent of modern fluorimetry, numerous in vitro studies employing primarily isolated mouse islets have investigated the effects of various secretagogues on cytoplasmic calcium, predominantly in insulin-secreting beta cells. Due to technical limitations, insights of these studies are inherently limited to a rather small subpopulation of outermost cells. The results also seem to depend on various factors, like culture conditions and duration, and are not always easily reconcilable with findings in vivo. The main controversies regard the types ofcalcium oscillations, presence of calcium waves, and the level of synchronized activity. Here, we set out to combine the in situ acute mouse pancreas tissue slice preparation with noninvasive fluorescent calcium labeling and subsequent confocal laser scanning microscopy to shed new light on the existing controversies utilizing an innovative approach enabling the characterization of responses in many cells from all layers of islets. Our experiments reproducibly showed stable fast calcium oscillations on a sustained plateau rather than slow oscillations as the predominant type of response in acute tissue slices, and that calcium waves are the mechanistic substrate for synchronization of oscillations. We also found indirect evidencethat even a large amplitude calcium signal was not sufficient and thatmetabolic activation was necessary to ensure cell synchronization upon stimulation with glucose. Our novel method helped resolve existing controversies and showed the potential to help answer important physiological questions, making it one of the methods of choice for the foreseeable future.
Ključne besede: glucose, pancreas, islets of Langerhans, mice
Objavljeno: 10.07.2015; Ogledov: 485; Prenosov: 178
.pdf Celotno besedilo (2,83 MB)
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2.
Novel insights into the downstream pathways and targets controlled by transcription factors CREM in the testis
Rok Košir, Peter Juvan, Martina Perše, Tomaž Büdefeld, Gregor Majdič, Martina Fink, Paolo Sassone-Corsi, Damjana Rozman, 2012, izvirni znanstveni članek

Opis: The essential role of the Crem gene in normal sperm development is widely accepted and is confirmed by azoospermia in male mice lacking the Crem gene. The exact number of genes affected by Crem absence is not known, however a large difference has been observed recently between the estimated number of differentially expressed genes found in Crem knock-out (KO) mice compared to the number of gene loci bound by CREM. We therefore re-examined global gene expression in male mice lacking the Crem gene using whole genome transcriptomeanalysis with Affymetrix microarrays and compared the lists of differentially expressed genes from Crem-/- mice to a dataset of genes where binding of CREM was determined by Chip-seq. We determined the global effect ofCREM on spermatogenesis as well as distinguished between primary and secondary effects of the CREM absence. We demonstrated that the absence of Crem deregulates over 4700 genes in KO testis. Among them are 101 genes associated with spermatogenesis 41 of which are bound by CREM and are deregulated in Crem KO testis. Absence of several of these genes in mouse models has proven their importance for normal spermatogenesis and male fertility. Our study showed that the absence of Crem plays a more important role on different aspects of spermatogenesis as estimated previously, with itsimpact ranging from apoptosis induction to deregulation of major circadian clock genes, steroidogenesis and the cell-cell junction dynamics. Several new genes important for normal spermatogenesis and fertility are down-regulated inKO testis and are therefore possible novel targets of CREM.
Ključne besede: spermatogenesis, genetics, Crem, mice
Objavljeno: 19.06.2017; Ogledov: 222; Prenosov: 151
.pdf Celotno besedilo (1,77 MB)
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3.
The relationship between membrane potential and calcium dynamics in glucose-stimulated beta cell syncytium in acute mouse pancreas tissue slices
Jurij Dolenšek, Andraž Stožer, Maša Skelin, Evan Miller, Marjan Rupnik, 2013, izvirni znanstveni članek

Opis: Oscillatory electrical activity is regarded as a hallmark of the pancreatic beta cell glucose-dependent excitability pattern. Electrophysiologically recorded membrane potential oscillations in beta cells are associated with in-phase oscillatory cytosolic calcium activity ([Ca2+]i) measured with fluorescent probes. Recent high spatial and temporal resolution confocal imaging revealed that glucose stimulation of beta cells in intact islets within acute tissue slices produces a [Ca2+]i change with initial transient phase followed by a plateau phase with highly synchronized [Ca2+]i oscillations. Here, we aimed to correlate the plateau [Ca2+]i oscillations with the oscillations of membrane potential using patch-clamp and for the first time high resolution voltage-sensitive dye based confocal imaging. Our results demonstrated that the glucose-evoked membrane potential oscillations spread over the islet in a wave-like manner, their durations and wave velocities being comparable to the ones for [Ca2+]i oscillations and waves. High temporal resolution simultaneous records of membrane potential and [Ca2+]i confirmed tight but nevertheless limited coupling of the two processes, with membrane depolarization preceding the [Ca2+]i increase. The potassium channel blocker tetraethylammonium increased the velocity at which oscillations advanced over the islet by several-fold while, at the same time, emphasized differences in kinetics of the membrane potential and the [Ca2+]i. The combination of both imaging techniques provides a powerful tool that will help us attain deeper knowledge of the beta cell network.
Ključne besede: glucose, pancreas, mice
Objavljeno: 19.06.2017; Ogledov: 376; Prenosov: 146
.pdf Celotno besedilo (2,57 MB)
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4.
Haploinsufficiency for steroidogenic factor 1 affects maternal behavior in mice
Tanja Španić, Neža Grgurevič, Gregor Majdič, 2016, izvirni znanstveni članek

Opis: Steroidogenic factor 1 (SF-1), officially designated NR5A1, is essential for gonadal and adrenal development and for the normal structure of the ventromedial hypothalamus (VMH), as demonstrated by SF-1 knockout mice (SF-1 KO), but much less is known about the possible effects of haploinsufficiency of the SF-1 gene. In the present study, maternal behavior in SF-1 KO heterozygous mice was evaluated. Behavioral tests revealed that SF-1 KO heterozygous females have impaired maternal behavior. In comparison to wild-type (WT) females, SF-1 KO heterozygous females retrieved significantly fewer pups into their nests, latency to retrieve and crouch over the pups was longer, and their nests were lower quality. As suggested by previous studies full dosage of SF-1 gene is needed for appropriate stress response and expression of brain-derived neurotrophic factor (BDNF) in the brain, and this might present a mechanism through which maternal behavior in SF-1 KO heterozygous females is impaired.
Ključne besede: mice, stress, steroidogenic factor 1, maternal behavior
Objavljeno: 10.07.2017; Ogledov: 261; Prenosov: 149
.pdf Celotno besedilo (477,98 KB)
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5.
Social isolation during puberty affects female sexual behavior in mice
Jasmina Kerčmar, Stuart Tobet, Gregor Majdič, 2014, izvirni znanstveni članek

Opis: Exposure to stress during puberty can lead to long-term behavioral alterations in adult rodents coincident with sex steroid hormone-dependent brain remodeling and reorganization. Social isolation is a stress for social animals like mice, but little is known about the effects of such stress during adolescence on later reproductive behaviors. The present study examined sexual behavior of ovariectomized, estradiol and progesterone primed female mice that were individually housed from 25 days of age until testing at approximately 95 days, or individually housed from day 25 until day 60 (during puberty), followed by housing in social groups. Mice in these isolated groups were compared to females that were group housed throughout the experiment. Receptive sexual behaviors of females and behaviors of stimulus males were recorded. Females housed in social groups displayed greater levels of receptive behaviors in comparison to both socially isolated groups. Namely, social females had higher lordosis quotients (LQs) and more often displayed stronger lordosis postures in comparison to isolated females. No differences between female groups were observed in stimulus male sexual behavior suggesting that female “attractiveness” was not affected by their social isolation. Females housed in social groups had fewer cells containing immunoreactive estrogen receptor (ER) α in the anteroventral periventricular nucleus (AVPV) and in the ventromedial nucleus of the hypothalamus (VMH) than both isolated groups. These results suggest that isolation during adolescence affects female sexual behavior and re-socialization for 1 month in adulthood is insufficient to rescue lordosis behavior from the effects of social isolation during the pubertal period.
Ključne besede: mice, social isolation, female sexual behavior, estrogen receptor α, puberty, adolescence
Objavljeno: 10.07.2017; Ogledov: 405; Prenosov: 204
.pdf Celotno besedilo (944,61 KB)
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