Concentration-dependence of dithiothreitol effects on rat distal colon electrophysiology

Dithiothreitol (DTT), at 1 mmol/L or higher, is widely used as a mucolytic in gastrointestinal research. Previous observations suggest that it may be toxic to the mucosa. DTT effects on the mucosal electrical behavior were assessed. Cumulative concentration-response relationships of DTT effects on rat distal colon mucosa were studied. Isolated mucosa preparations were mounted in an Ussing chamber under short-circuit conditions. The effects of concentrations ranging from 1 mumol/L to 1 mmol/L, applied to either the mucosal or serosal side, were studied. As compared with control, untreated preparations, DTT depressed short-circuit current at 10 mumol/L and higher when applied to the serosal side, and at 50 mumol/L and higher when applied to the mucosal side of the epithelium. On the other hand, transepithelial resistivity showed a progressive increase with DTT applied to either side at a concentration of up to 500 mumol/L, while at the highest concentration (1 mmol/L) a marked decrease in resistivity occurred. Neither the short-circuit current decrease, nor the resistivity collapse showed recovery after repeated rinsing with DTT-free solution. It is concluded that DTT affects epithelial electrical properties at low concentrations, and therefore its use as a mucolytic for electrophysiological studies should be discouraged.

Sodium-deprived rat distal colon epithelial response to acute hypoxia and reoxygenation

In normal rat distal colon isolated mucosa, basal short-circuit current (Isc) is mostly due to chloride secretion. Isc is depressed by a brief (5 min) acute hypoxia and overshoots above baseline during reoxygenation. Sodium deprivation raises serum aldosterone levels and leads to expression of functional epithelial sodium channels which are amiloride-sensitive. Thus, in sodium-deprived rats (SDRs) Isc is dependent on electrogenic sodium absorption. Since the ion primarily responsible for the Isc is different in each functional condition, it is not known whether hypoxia and reoxygenation affect SDRs epithelial response in the same way as in normal rats. Therefore the electrical behavior of isolated mucosa preparations from normal and SDRs was studied in an Ussing chamber, and the effect of the epithelial sodium channel blocker, amiloride sensitive, basal Isc than controls. Their response to hypoxia (expressed as a fraction of basal Isc) was similar to controls but upon reoxygenation their recovery was incomplete. SDRs response to hypoxia was not affected by amiloride at any concentration tested. However, post-hypoxic recovery was modified by amiloride in a concentration-dependent way: it was incomplete at 10(-8) M, complete at 10(-6) M, and at 10(-4) M it overshooted above baseline values. Therefore, in sodium-deprived rats, sodium channel blockade reverts the pattern of blunted recovery to the overshooting pattern seen normal rats. These results may be explained by two non-mutually exclusive hypotheses: Epithelial sodium channel blockade in sodium-deprived rats might (1) unmask a basal chloride conductance, and (2) interfere with a negative interaction between sodium chloride conductances.

Rat colon epithelium response to hypoxia is modified by intrinsic innervation blockade

Background/Aim: Short-circuit current (Isc) and transepithelial potential difference (PD) of rat distal colon decrease during acute hypoxia and overshoot on reoxygenation. It is not known whether tonic intrinsic nervous activity may influence these responses. Methods: Preparations lacking the submucosal plexus (isolet mucosa) and preparations retaining it (mucosa-submucosa) were mounted in Ussing chambers at 37 degrees Celsius and gassed with 95 percent O2 -5 percent CO2; Isc and PD were monitored. A 5-min hypoxia with 95 percent N2-5 percent CO2 was followed by reoxygenation. The procedure was repeated in the presence of the nervous blocking agent, tetrodotoxin (10(-6)M) in the serosal side of the chamber. Results: In the isolated mucosa (n=10) hypoxia reduced Isc by -55 + 5 percent and PD by -54 + 6 percent below baseline; reoxygenatory overschoots were, respectively, + 60 + 17 percent and + 16 percent. Tetrodotoxin slightly and transiently reduced baseline Isc (-16 + 2 percent) and PD (-14 + 3 percent), with a small resistivity increase. It did not significatively modify the responses to responses to either hypoxia or reoxygenation. In mucosa-submucosa preparations (n=9) hypoxia reduced Isc (-54 + 8 percent) and PD (-61 + 4 percent). On reoxygenation Isc and PD were increased, respectively, +30 + 5 percent and +19 + 6 percent over baseline. Tetrodotoxin reduced baseline Isc (-59,6 + 5 percent) and PD (61,3 + 6 percent). It enhanced hypoxic Isc and PD decreases (-80 + 5 percent), but not the reoxygenatory overschoots. Conclusions: 1) Tetrodotoxin affects baseline Isc and PD more intensely in submucosal plexus innervated preparations than in the isolated mucosa. 2) The epithelial electrical response to acute hypoxia appears to be modulated by tonic neural activity.

RM del SNC: uso de una secuencia inversión recuperación sensible al edema / CNS RM imaging: use of an inversion recovery pulse sequence for brain edema

El edema cerebral es el denominador común de un amplio espectro de patologías que frecuentemente afectan al Sistema Nervioso Central (SNC). En la actualidad, su diagnóstico y fundamentalmente su delimitación resultan difíciles con las consecuencias spin-eco (SE de uso habitual en las imágenes por Resonancia Magnética). Con el objeto de superar este inconveniente utilizamos una secuencia Inversión Recuperación (IR), con tiempo de repetición (TR), tiempo de inversión (TI), y tiempo de eco (TE) largos, desarrollada para tal fin. El análisis de la imágenes obtenidas muestra al líquido cefalorraquídeo (LCR) con señal fuertemente hipointensa, el cerebro normal con señal intermedia, las zonas afectadas por tumores u otras patologías con señal hipointensa en relación al cerebro, y el edema como el único elemento hiperintenso permitiendo una clara diferenciación del mismo, con elevada relación contraste-ruido (C/R)

Colon epithelial electrical responses to acute hyposia and reoxygenation in vitro

Electrogenic epithelial transport depends on oxidative metabolism. Acute hypoxia and subsequent reoxygenation effects on short-circuit current (Isc), transepithelial potential difference (PD) and tissue resistivity (TR) of rat distal colon were assessed. The tissue was mounted in an Ussing chamber filled with Ringer-HCO3 -solution at 37 degrees Celsius and bubbled with 95 per cent O2- 5 per cent CO2 which was switched to 95 per cent N2- 5 per cent CO2 for inducing hypoxia; afterwards normal oxygenation was resumed. The effect of 5, 10, 15 and 20 min-hypoxic periods was assessed in isolataed mucosa preparations. Recovery was complete after 10- and 15-min hypoxia, but not after 20-min hypoxia. After 5-min hypoxia, an overshoot of Isc and PD was seen on reoxygenation. This effect was further characterized comparatively in mucosa-submucosa and isolated mucosa preparations. In the former (n=10), control values were Isc= 71,7 + 8,6 muA,. cm-2, PD = 9,7 + 1,6 mV and TR = 134,9 + 13,6 (. cm2. A 5-min hypoxia reduced Isc by 47.2 + 7,3 per cen+t and PD by 61,5 + 4,9 per cent. Peak values on reoxygenation were 28,1 + 4,1 per cent for Isc and 16,8 + 5,4 per cent for PD, over controls values. In the osilated mucosa (n=9), control values were Isc= 52,04 + 5,5 muA. cm-2, PD = 5,0 + 0,8 mV and TR = 101,04 + 10,5 (. cm2. In hypoxia, Isc decreased by 64,5 + 7,6 per cent and PD by 57,2 + 7,8 per cent. On reoxygenation peak values of 78,0 + 19,0 per cent and 87,5 + 17,1 per cent, respectively, were seen. The response to a 5 min-hypoxia was comparable, but that to reoxygenation was weaker and slower, in mucosa-submucosa than in isolated mucosa preparations. This may be explained by a hindrance to oxygen diffusion caused by the submucosal tissue. TR did not change with any period of hypoxia tested, but decreased slightly (8,9 + 1,3 per cent) upon reoxygenation in the mucosa-submucosa preparations. Ouabain (10(-3)M) markedly blunted the response to reoxygenation. We conclude that hypoxic periods of 20 min lead to irreversible functional deterioration. Hypoxia decreases electrogenic transepithelial pumping, which may allow sodium to accumulate intracellularly and, if the hypoxia is short enough to prevent damage to the epithelium, increase sodium pump activity when oxygenation is resumed.