The chloride anion as a signalling effector.

The specific role of the chloride anion (Cl- ) as a signalling effector or second messenger has been increasingly recognized in recent years. It could represent a key factor in the regulation of cellular homeostasis. Changes in intracellular Cl- concentration affect diverse cellular functions such as gene and protein expression and activities, post-translational modifications of proteins, cellular volume, cell cycle, cell proliferation and differentiation, membrane potential, reactive oxygen species levels, and intracellular/extracellular pH. Cl- also modulates functions in different organelles, including endosomes, phagosomes, lysosomes, endoplasmic reticulum, and mitochondria. A better knowledge of Cl- signalling could help in understanding the molecular and metabolic changes seen in pathologies with altered Cl- transport or under physiological conditions. Here we review relevant evidence supporting the role of Cl- as a signalling effector.

Cloreto e status hidroeletrolítico e ácido-básico: importância clínica da determinação laboratorial / Chloride and hydroelectrolytic and acid base status: clinical importance of laboratorial determination

Apesar de o cloreto ser um exame amplamente disponível e solicitado em instituições de saúde, a maior pane dos médicos crê ser difícil interpretar seus resultados, estabelecer correlações com outros parâmetros laboratoriais associados e tomar decisões terapêuticas baseadas no mesmo. Devido à complexidade de sua homeostase e à intrincada correlação com o status hidroeletrolítico e ácido-básico, poucos médicos se sentem efetivamente aptos a aproveitar a valiosa informação clínica que ele pode revelar. Isto é agravado pela rarefeita bibliografia objetiva sobre o tema. Os autores desta revisão não foram capazes de encontrar nenhum capítulo especificamente dedicado ao cloreto nos principais livros-texto de Fisiologia, Clínica Médica e Nefrologia, disponíveis, à exceção de três revisões bibliográficas no Medline. Não obstante, tentamos organizar a informação tão claramente quanto possível, com o objetivo de tornar o cloreto uma ferramenta útil aos nossos colegas profissionais de saúde.

Even though chloride is a widely available and requested test in health institutions, most part of physicians find it difficult to interpret its results, establish correlations with other laboratory linked parameters and take therapeutic decisions based on it. Due to the complexity of its homeostatic balance and intrincated correlation to hydroelectrolytic and acid base status, few doctors feel actually able to fully profit from the valuable clinical information it can unfold. This is aggravated by the scarce objective bibliography on the issue. The authors of this review were not able to find any chapters specifically dedicated to chloride on major Physiology, Internal Medicine and Nephrology textbooks, but only three reviews on Medline. Nevertheless, we managed to organize the information as clearly as possible with the aim of making chloride test an useful tool to our fellow health professionals.

Effects of androgen manipulation on alpha1-adrenoceptor subtypes in the rat seminal vesicle.

This study analyses possible changes during surgical and chemical castration in the expression and pharmacological characteristics of alpha1-adrenoceptor in adult rat seminal vesicle. Ribonuclease protection assays indicated that alpha1a--was the predominant mRNA, while alpha1b-and alpha1d-adrenoceptor transcripts were detected in lower abundance in this tissue. alpha1a-adrenoceptor mRNA expression presented a complex dependency on androgens, while alpha1b- and alpha1d-adrenoceptor transcripts were both upregulated with surgical and chemical castration, suggesting a negative modulation by androgens. Testosterone treatment reversed the effects caused by surgical castration. Functional studies confirmed the involvement of alpha1A- and alpha1B-adrenoceptor in the seminal vesicle contractile responses, and suggested that alpha1B-induced contractile response was upregulated after castration. Taken together, the results suggest that alpha1-adrenoceptor expression in seminal vesicle is differentially regulated by the androgen status of the rat.

Effects of intracellular calcium elevation on action potential and L-type calcium current of normal and chronically infarcted rat ventricles.

The present work investigated the effects of raising [Ca+2]i levels on action potential (AP) and L-type calcium current (I(Ca.L)) of normal and chronically infarcted rat ventricles. Experiments were performed by conventional electrophysiology and whole-cell patch-clamp techniques. In the former, APs were recorded in ventricular strips subjected to different pacing rates or elevation of [Ca+2]o levels. In the latter, I(Ca.L) was studied in isolated myocytes in the absence of an intracellular Ca+2 chelator. The acceleration of heart rate (6 to 240 beats/min) reduced AP duration measured at 20%, 50%, and 90% repolarization (APD20, APD50, and APD90) in the infarcted group, and increased APD20 and APD50 in the control group. Rising [Ca+]o (1.25 to 5.0 mmol/L) induced a decrease of APD20 and APD50 in both groups. Voltage clamp revealed a smaller I(Ca.L) density at approximately -17 mV in myocytes from infarcted ventricles (-1.86 +/- 0.37 vs -3.98 +/- 0.65 pA/pF, P < .05), and the appearance of a non-K+ outward current coupled to I(Ca.L). The results suggest the participation of a Ca+2-activated outward current in the repolarization of normal and infarcted rat ventricles.

Role of Cl- in electrogenic H+ secretion by cortical distal tubule.

The presence of an electrogenic H+-ATPase has been described in the late distal tubule, a segment which contains intercalated cells. The present paper studies the electrogenicity of this transport mechanism, which has been demonstrated in turtle bladder and in cortical collecting duct. Transepithelial PD (Vt) was measured by means of Ling-Gerard microelectrodes in late distal tubule of rat renal cortex during in vivo microperfusion. The tubules were perfused with electrolyte solutions to which 2 x 10(-7) M bafilomycin or 4.6 x 10(-8) M concanamycin were added. No significant increase in lumen-negative Vt upon perfusion with these inhibitors as compared to control, was observed as well as when 10(-3) m amiloride, 10(-5) M benzamil or 3 mM Ba2+ were perfused alone or in combination. The effect of an inhibition of electrogenic H+ secretion, i.e., increase in lumen-negative Vt by 2-4 mV, was observed only when Cl- channels were blocked by 10(-5) M 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). This blocker also reduced the rate of bicarbonate reabsorption in this segment from 1.21 +/- 0.14 (n = 8) to 0.62 +/- 0.03 (8) nmol.cm-2.sec-1 as determined by stationary microperfusion and pH measurement by ion-exchange resin microelectrodes. These results indicate that: (i) the participation of the vacuolar H+ ATPase in the establishment of cortical late distal tubule Vt is minor in physiological conditions, but can be demonstrated after blocking Cl- channels, thus suggesting a shunting effect of this anion; and, (ii) the rate of H+ secretion in this segment is reduced by a Cl- channel blocker, supporting coupling of H+-ATPase with Cl- transport.

Dopamine transporter mediated release of dopamine: role of chloride.

Using a rapid (0.5 ml/min) flow rate superfusion system, the dopamine (DA) transporter mediated release of DA is further explored, and compared to the depolarization evoked release of DA in rat striatal synaptosomes preloaded with radioactive DA (3H-DA). In this system external DA in the low microM range efficaciously releases the preloaded transmitter, the maximal response being reached at 3 microM DA. The external DA stimulated release is Ca(2+)-independent, Cl(-)-dependent, and blocked by both bupropion and nomifensine. The atypical antidepressant bupropion inhibits 3H-DA accumulation to rat striatal synaptosomes with a calculated IC50 of 1.3 x 10(-6) M. Among DA uptake blockers some are known to act as DA releasing agents. Here we found that the DA uptake blocker nomifensine (30 microM) is unable to modify the baseline release of 3H-DA, whereas bupropion (10 microM) clearly elevates the baseline release of 3H-DA in a Ca(2+)-independent and Cl(-)-dependent manner. The non releasing agent nomifensine blocks the release of 3H-DA induced by bupropion. The Ca(2+)-dependent, high K+ depolarization evoked release of 3H-DA is not modified by nomifensine and does not depend on the external Cl- concentration. When the depolarizing medium contains DA the carrier mediated release of 3H-DA induced by the external DA is additive to the high K+ induced response. A drastic drop in the external Cl- concentration induces 3H-DA release. This release of 3H-DA induced by low external Cl- levels is completely blocked by nomifensine, which only slightly diminished the release of 3H-DA induced by the absence of external Na+. On the basis of these results, it is concluded that: 1) Rapid perfusion flow rates eliminate DA reuptake. 2) DA uptake inhibitors either with or without DA releasing capabilities block the release of DA induced by microM levels of external DA. 3) By preventing translocation of the DA transporter mobile moiety, nomifensine may inhibit the release of DA induced by external DA or bupropion and by drastic drops in the external Cl- concentration. 4) In the absence of nomifensine, the DA transporter works under both resting and depolarized conditions, but in contrast to the GABA transporter (Sitges et al.: Neurochem Res 18:1081-1087, 1993), the DA transporter does not contribute to the amount of the DA released by depolarization. 5) Reversal of the DA uptake carrier is favored by conditions increasing the internal DA levels. 6) Cl- rather than Na+ is a major determinant in 3H-DA movements through the DA transporter.

Differential effects of GABAergic ligands in mouse and rat hippocampal neurons.

Previous electrophysiological studies suggested that GABAA receptors in rat hippocampal neurons might be less sensitive to ethanol than mouse neurons. Therefore, we examined the effects of ethanol (0.5-850 mM) in cultured mouse (C57BL/6) and rat (Sprague-Dawley) neurons. In 35% of the mouse neurons, the Cl- current was potentiated by ethanol starting at 0.5 mM. In all of the rat neurons examined, on the other hand, the current was potentiated by concentrations starting at 200 mM. We also studied the effects of GABA and other GABAergic ligands. GABAA receptors in rat and mouse neurons displayed EC50s for GABA of 9 +/- 0.3 and 17 +/- 0.8 microM, respectively and ethanol did not significantly change these values. The EC50 for diazepam was 92 +/- 3 and 120 +/- 8 nM in rat and mouse, respectively. Pentobarbital enhanced the current with EC50s of 84 +/- 3 and 106 +/- 6 microM in rat and mouse, respectively. The sensitivity for Cl-218,872, which binds preferentially to the Type I benzodiazepine receptor, was similar in all the neurons. RO 15-4513, an inverse partial agonist to the benzodiazepine receptor, was not effective in reversing the potentiation of the Cl- current in rat neurons and only slightly reduced the potentiation in mouse neurons. The receptors in rat neurons were more sensitive to external Zn2+; the current was inhibited by 50% with a concentration of 93 +/- 3 and 244 +/- 9 microM in rat and mouse, respectively. Analysis of mRNA encoding for the gamma 2L receptor subunit showed similar levels in rat and mouse neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Secretory effect of serotonin on jejunal response to cholera toxin, in dog

A substância mediadora liberada pela toxina da cólera e que estimula a secreçäo intestinal é ainda desconhecida. Sabe-se que a serotonina está envolvida no estímulo secretor intestinal de água e eletrólitos. Tendo em vista a avaliaçäo de um provável papel da serotonina na induçäo secretora jejunal pela toxina da cólera, calcularam-se os volumes de água, sódio, potássio e cloreto, bem como os níveis imunorreativos de serotonina, em alça de Thiry-Vella canina. A administraçäo de toxina provocou um aumento na secreçäo de todos os eletrólitos e do fluxo de serotonina. Esses resultados sugerem que a toxina da cólera induz à liberaçäo de serotonina na luz intestinal, talvez como um mediador da secreçäo hidroeletrolítica entérica

Second messenger-mediated actions of norepinephrine on target neurons in central circuits: a new perspective on intracellular mechanisms and functional consequences.

Ever since the initial demonstration of a widespread distribution of noradrenergic fibers to functionally diverse regions of the mammalian forebrain, there has been considerable interest in determining the electrophysiological effects of norepinephrine (NE) on individual neurons within these target areas. While early studies showed that NE could directly inhibit cell firing via increased intracellular levels of cyclic AMP, more recent work has revealed a spectrum of noradrenergic actions, which are more accurately characterized as neuromodulatory. More specifically, numerous experimental conditions have been described where NE at levels subthreshold for producing direct depressant effects on spontaneous firing can facilitate neuronal responses to both excitatory and inhibitory synaptic stimuli. The goal of this report is to review recent evidence which suggests that the various modulatory actions of NE on central neurons result from the activation of different adrenoceptor-linked second messenger systems. In particular, we have focused on the candidate signal transduction mechanisms that may underlie NE's ability to augment cerebellar and cortical neuronal responsiveness to GABAergic synaptic inputs. The consequences of such NE-induced changes in synaptic efficacy are considered not only with respect to their influences on feature extraction properties of individual sensory cortical neurons but also with regard to the potential impact such actions would have on the signal processing capabilities of a network of noradrenergically innervated cortical cells.

Calcium-dependent chloride currents elicited by injection of ethanol into Xenopus oocytes.

Electrophysiological techniques were used to study the response of native Xenopus laevis oocytes to intracellular injection and bath application of ethanol. Injection of ethanol produced dose-dependent transient inward currents accompanied by large current fluctuations, with estimated intracellular concentrations ranging from 10 to 300 mM. The response duration varied between 2 and 15 min, with an onset delay of 2-15 s. The inward current sometimes consisted of a fast and a slow component. Bath application of equivalent concentrations elicited similar but considerably smaller responses. The current showed a reversal potential of -20 +/- 10 mV, corresponding to an increase in chloride permeability. The response was eliminated in the presence of low chloride saline and was blocked by the chloride channel inhibitors SITS and DIDS. Ethanol responses were inhibited by the intracellular injection of the calcium chelator EGTA and were unaffected when the extracellular calcium was lowered. It is concluded that ethanol injection into Xenopus oocytes elicits a release of calcium from intracellular stores, which then activates an increased membrane permeability to chloride.

Intracellular chloride regulation in amphibian dorsal root ganglion neurones studied with ion-selective microelectrodes.

1. Intracellular Cl- activity (aiCl) and membrane potential (Em) were measured in frog dorsal root ganglion neurones (DRG neurones) using double-barrelled Cl- -selective microelectrodes. In standard Ringer solution buffered with HEPES (5 mM), equilibrated with air or 100% O2, the resting membrane potential was -57.7 +/- 1.0 mV and aiCl was 23.6 +/- 1.0 mM (n = 53). The value of aiCl was 2.6 times the activity expected for an equilibrium distribution and the difference between Em and ECl was 25 mV. 2. Removal of external Cl- led to a reversible fall in aiCl. Initial rates of decay and recovery of aiCl were 4.1 and 3.3 mM min-1, respectively. During the recovery of aiCl following return to standard Ringer solution, most of the movement of Cl- occurred against the driving force for a passive distribution. Changes in aiCl were not associated with changes in Em. Chloride fluxes estimated from initial rates of change in aiCl when external Cl- was removed were too high to be accounted for by electrodiffusion. 3. The intracellular accumulation of Cl- was dependent on the extracellular Cl- activity (aoCl). The relationship between aiCl and aoCl had a sigmoidal shape with a half-maximal activation of about 50 mM-external Cl-. 4. The steady-state aiCl depended on the simultaneous presence of extracellular Na+ and K+. Similarly, the active reaccumulation of Cl- after intracellular Cl- depletion was abolished in the absence of either Na+ or K+ in the bathing solution. 5. The reaccumulation of Cl- was inhibited by furosemide (0.5-1 x 10(-3) M) or bumetanide (10(-5) M). The decrease in aiCl observed in Cl- -free solutions was also inhibited by bumetanide. 6. Cell volume changes were calculated from the observed changes in aiCl. Cells were estimated to shrink in Cl- -free solutions to about 75% their initial volume, at an initial rate of 6% min-1. 7. The present results provide direct evidence for the active accumulation of Cl- in DRG neurones. The mechanism of Cl- transport is electrically silent, dependent on the simultaneous presence of external Cl-, Na+ and K+ and inhibited by loop diuretics. It is suggested that a Na+:K+:Cl- co-transport system mediates the active transport of Cl- across the cell membrane of DRG neurones.

Active transport of the chloride ion by the colon - abstract

The present data represent some results of our continuing studies on the mechanisms of electrolyte transport in the colon. Trained dogs with chronic isolated segments of colon were used, and the transport of chloride ion was studied by instilling the following electrolyte solutions into the colon: sodium chloride (154 mEq/L, 100 mEq/L), Mannitol (5 percent) choline choride (100 mEq/L), lithium chloride (100 mEq/L), lithium chloride (60 mEq/L) plus potassium chloride (40 mEq/L), lithium chloride (75 mEq/L) plus sodium bicarbonate (75 m Eq/L). All solutions were made isotonic whenever necessary by the addition of mannitol and all contained the non-absorbable maker polyethylene glycol (PEG). Samples of colonic fluid were taken at intervals over a six-hour period following the instillation of solution into the isolated colonic segment. The samples were analysed for Na, K, Cl, HCO3 - pH, osmolality and PEG. The electrical potential difference across the colon was measured in several experiments. Chloride ion transport was also studied following the administration of Diamax or Aldosterone, mucosa. There was no evidence of active water transport. The mucosa is normally electrically negative with respect to serosa; however, in the presence of choline chloride the mucosa becomes electrically positive with respect to serosa. Our data also show that the chloride ion is actively transported against both an electrical and a chemical gradient. In addition chloride can be transported in the absence of sodium, but in the presence of sodium can be transported more rapidly than that ion. Chloride transport is to some extent linked to sodium transport and also to bicarbonate transport (AU)