Neuronal activity and many pathological states in the CNS are accompanied by transient astrocytic swelling, which affects excitability, extrasynaptic transmission, and neuron-glia interactions. By using three-dimensional confocal morphometry (3DCM), we quantified the morphometric parameters of astrocytes in intact tissue. In experiments performed in brain cortex slices from transgenic GFAP/EGFP mice, we applied 3DCM to study the dynamic changes in astrocyte morphology during hypotonic stress. Our morphometric analysis showed that the effect of a 10-min application of hypotonic solution (200 mmol/kg) on the swelling of different cell compartments was dependent on the extent of the swelling of the total astrocyte volume. If the swelling of the whole cell, i.e., soma and processes, was less than approximately 10%, there were no differences between the swelling of the soma and the processes. However, if the swelling of the total cell volume was greater than 10%, the swelling of the processes was greater than the swelling of the soma. Analyzing the effect of hypotonic solution on the morphology of these astrocytes revealed that the total cell volume increased; however, certain cell compartments were distinguished in which the volume increased, whereas in other compartments cell volume decreased or apparently did not change, and the structure of some compartments was altered. Our data show that astrocytes in brain slices undergoing hypotonic stress display cell volume regulation as well as transient changes in morphology.

Energy depletion during ischemia leads to disturbed ionic homeostasis and accumulation of neuroactive substances in the extracellular space, subsequently leading to volume changes in astrocytes. Confocal microscopy combined with 3D reconstruction was used to quantify ischemia-induced astrocyte volume changes in cortical slices of GFAP/EGFP transgenic mice. Twenty-minutes of oxygen-glucose deprivation (OGD) or oxygen-glucose deprivation combined with acidification (OGD(pH 6.8)) revealed the presence of two distinct astrocytic populations, the first showing a large volume increase (HR astrocytes) and the second displaying a small volume increase (LR astrocytes). In addition, changes in resting membrane potential (V(m)), measured by the patch-clamp technique, supported the existence of two astrocytic populations responding differently to ischemia. Although one group markedly depolarized during OGD or OGD(pH 6.8), only small changes in V(m) toward more negative values were observed in the second group. Conversely, acidification (ACF(pH 6.8)) led to a uniform volume decrease in all astrocytes, accompanied by only a small depolarization. Interestingly, two differently responding populations were not detected during acidification. Differences in the expression of inwardly rectifying potassium channels (Kir4.1), glial fibrillary acidic protein (GFAP), and taurine levels in cortical astrocytes were detected using immunohistochemical methods. We conclude that two distinct populations of astrocytes are present in the cortex of GFAP/EGFP mice, based on volume and V(m) changes during exposure to OGD or OGD(pH 6.8). Immunohistochemical analysis suggests that the diverse expression of Kir4.1 channels and GFAP as well as differences in the accumulation of taurine might contribute to the distinct ability of astrocytes to regulate their volume.

Soták M, Polidarová L, Musílková J, Hock M, Sumová A, Pácha J. Circadian regulation of electrolyte absorption in the rat colon. Am J Physiol Gastrointest Liver Physiol 301: G1066-G1074, 2011. First published September 8, 2011 doi:10.1152/ajpgi.00256.2011The intestinal transport of nutrients exhibits distinct diurnal rhythmicity, and the enterocytes harbor a circadian clock. However, temporal regulation of the genes involved in colonic ion transport, i.e., ion transporters and channels operating in absorption and secretion, remains poorly understood. To address this issue, we assessed the 24-h profiles of expression of genes encoding the sodium pump (subunits Atp1a1 and Atp1b1), channels (␣-, ␤-, and ␥-subunits of Enac and Cftr), transporters (Dra, Ae1, Nkcc1, Kcc1, and Nhe3), and the Na ϩ /H ϩ exchanger (NHE) regulatory factor (Nherf1) in rat colonic mucosa. Furthermore, we investigated temporal changes in the spatial localization of the clock genes Per1, Per2, and Bmal1 and the genes encoding ion transporters and channels along the crypt axis. In rats fed ad libitum, the expression of Atp1a1, ␥Enac, Dra, Ae1, Nhe3, and Nherf1 showed circadian variation with maximal expression at circadian time 12, i.e., at the beginning of the subjective night. The peak ␥Enac expression coincided with the rise in plasma aldosterone. Restricted feeding phase advanced the expression of Dra, Ae1, Nherf, and ␥Enac and decreased expression of Atp1a1. The genes Atp1b1, Cftr, ␣Enac, ␤Enac, Nkcc1, and Kcc1 did not show any diurnal variations in mRNA levels. A low-salt diet upregulated the expression of ␤Enac and ␥Enac during the subjective night but did not affect expression of ␣Enac. Similarly, colonic electrogenic Na ϩ transport was much higher during the subjective night than the subjective day. These findings indicate that the transporters and channels operating in NaCl absorption undergo diurnal regulation and suggest a role of an intestinal clock in the coordination of colonic NaCl absorption. rhythm; intestinal transport; transporters; channels VARIOUS FUNCTIONS OF THE INTESTINE such as motility, epithelial cell proliferation, digestion, and transport exhibit diurnal activities (9,13,30). Several transporters involved in absorption of carbohydrates and small peptides exhibit robust circadian fluctuations that seem to be regulated by two pathways. The first pathway involves food intake and is mediated by gut luminal signals, whereas the second one involves systemic entrainment pathways (21,22,35,40). In addition to nutrient absorption in the small intestine, the colonic absorption of some electrolytes and volatile fatty acids (5, 41, 42) and some intestinal secretory activity (4, 31, 32) exhibit diurnal rhythmicity.Functional circadian clocks operate in both the small (6, 23, 25) and large intestine (10,33,24). The molecular clock mechanism is based on interaction between transcription-based feedback loops. The transcription factors CLOCK and BMAL1 drive the transcription of Per, Cry, Rev-erb␣, and Ror␣ clock genes by binding CLO...