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Biophysical characterization of two mycobacteriophages (Phlei phage and Butyricum phage) was carried out. Biophysical parameters obtained were: (i) buoyant densities in CsCl of I . 51 g/ml for both phages; (ii) sedimentation coefficient of 490 S and 410 S; (iii) DNA content of 42 and 34% and (iv) mol. wt. calculated by electron microscopic dimensions to be 123 x 10(6) and 116 x 10(6) for Butyricum and Phlei phage, respectively.
Electron paramagnetic resonance (EPR) is a spectroscopic technique that allows detection of paramagnetic compounds, e.g., free radicals, in skin, and is used for measuring skin membrane fluidity and polarity. EPR imaging is concerned with spatially resolved EPR spectroscopy. We studied EPR images at X-band frequency (9 GHz) in the skin biopsies of hairless mice. Our particular imaging technique utilized a modulated field gradient to obtain cross-sectional images perpendicular to the skin surface. Employing nitroxide free radicals, this approach allows analysis of skin biophysical and biochemical features at the micrometer resolution level. By spin labeling drugs, pharmacokinetic properties of the labeled compound can be monitored in skin. We suggest that EPR imaging has a broad application potential in dermatologic research. In comparison to other spatially visualizing techniques, such as nuclear magnetic resonance (NMR) imaging and ultrasound, EPR imaging has significant advantages, such as high spatial resolution and providing specific biochemical and biophysical information that cannot be obtained by other methods in skin.
OBJECTIVE: To determine whether computer assisted fetal heart rate analysis or the biophysical profile score can provide noninvasive prediction of fetal acidaemia. DESIGN: Cross sectional study. SETTING: Harris Birthright Research Centre for Fetal Medicine, King's College Hospital School of Medicine, London. SUBJECTS: Forty-one women with pregnancies complicated by diabetes mellitus. INTERVENTIONS: Fetal heart rate (FHR) monitoring with computer assisted analysis, biophysical profile score (BPS) and cordocentesis for measurement of umbilical venous blood glucose concentration and blood gases, up to 24 h before delivery at 27 to 39 weeks gestation. RESULTS: The mean umbilical venous blood pH was significantly lower than the normal mean for gestation, and was below the 5th centile in 18 pregnancies, including all six cases where the mother had nephropathy and hypertension. The mean pO2 was not significantly different from the normal mean for gestation. There were significant associations between fetal acidaemia and both the BPS (r = 0.46, P < 0.01) and FHR variation (r = 0.42, P < 0.01). However, of the 12 acidaemic fetuses of non-nephropathic mothers, nine had normal BPS and six had normal FHR variation. CONCLUSIONS: In pregnancies complicated by maternal diabetes mellitus, BPS and FHR variation are of limited value in the prediction of fetal blood pH.
The subtle dryness of the skin surrounding the lesions of atopic dermatitis (AD) is called atopic dry skin or atopic xerosis (AX). AX is more susceptible to the development of AD skin lesions under various environmental stimuli than the clinically normal skin of the people who have or have had or will have AD, which might be called normal atopic skin (NAS) that shows no functional differences as compared to the skin of normal individuals. Routine histopathologic studies of AX that involve the invasive procedures of biopsy are not so helpful in clarifying the underlying pathogenesis. Modern, noninvasive biophysical instrumentation provides rich and quantitative information about various functional aspects of skin. The stratum corneum (SC) of AX reveals not only decreased hydration but also mildly impaired barrier function demonstrable as an increase in transepidermal water loss, elevated pH values, and an increased turnover rate of the SC consisting of thick layers of smaller-sized corneocytes. These data suggest that AX is related to mildly increased epidermal proliferation as a result of the presence of subclinical cutaneous inflammation. Although AX skin does not display any impairment in the recovery of barrier function after physical skin irritation by tape-stripping, it produces a much more severe, long-lasting inflammatory response together with a delay in barrier repair after chemical irritation such as that induced by sodium lauryl sulphate. The SC of AX is biochemically characterized by reduction in the amounts of ceramides, especially ceramide I, sebum lipids, and water-soluble amino acids. None of these changes in SC functions are seen in NAS, which includes not only the normal-looking skin of AD patients long after regression of all active lesions but also of latent atopic skin such as neonates who later develop AD. This suggests that all of the observed functional as well as biochemical abnormalities of AX are a reflection of subclinical inflammation. The presence of the underlying inflammation in AX also differentiates it from senile xerosis. The mildly impaired SC functions of AX can be improved by daily repeated applications of effective moisturizers, i.e., corneotherapy, which is effective in preventing the exacerbating progression of AX to AD resulting from inadvertent scratching of the skin that facilitates the penetration of environmental allergens into the skin. The biophysical confirmation of such efficacy of moisturizers, including cosmetic bases on the mildly impaired barrier function and decreased water-holding capacity of the SC of AX, definitely substantiates the importance of skin care for the cosmetic skin problems that affect every individual in the cold and dry season ranging from late autumn to early spring.
Synthetic rhamnolipids, derived from a natural diacylated glycolipid, RL-2,2(14), produced by Burkholderia (Pseudomonas) plantarii, were analyzed biophysically. Changes in the chemical structures comprised variations in the length, the stereochemistry and numbers of the lipid chains, numbers of rhamnoses, and the occurrence of charged or neutral groups. As relevant biophysical parameters, the gel (beta) to liquid crystalline (alpha) phase behavior of the acyl chains of the rhamnoses, their three-dimensional supramolecular aggregate structure, and the ability of the compounds to intercalate into phospholipid liposomes in the absence and presence of lipopolysaccharide-binding protein were monitored. Their biological activities were examined as the ability to induce cytokines in human mononuclear cells and to induce chemiluminescence in monocytes. Depending on the particular chemical structures, the physicochemical parameters as well as the biological test systems show large variations. This relates to the acyl chain fluidity, aggregate structure, and intercalation ability, as well as the bioactivity. Most importantly, the data extend our conformational concept of endotoxicity, based on the intercalation of naturally originating amphiphilic virulence factors into membranes from immune cells. This 'endotoxin conformation', produced by amphiphilic molecules with a hydrophilic charged backbone and apolar hydrophobic moiety, and adopting inverted cubic aggregate structures, causes high mechanical stress in target immune cells on integral proteins, eventually leading to cell activation. Furthermore, biologically inactive rhamnolipids with lamellar aggregate structures antagonize the endotoxin-induced activity in a way similar to lipid A-derived antagonists.
Before the onset of hearing, a transient efferent innervation is found on inner hair cells (IHCs). This synapse is inhibitory and mediated by a nicotinic cholinergic receptor (nAChR) probably formed by the alpha9 and alpha10 subunits. We analysed the pharmacological and biophysical characteristics of the native nAChR using whole-cell recordings from IHCs in acutely excised apical turns of the rat organ of Corti. Nicotine did not activate but rather blocked the acetylcholine (ACh)-evoked currents with an IC50 of 1 +/- 0.1 microM. Antagonists of non-cholinergic receptors such as strychnine, bicuculline and ICS-205930 blocked ACh-evoked responses with an IC50 of 8.6 +/- 0.8 nM, 59 +/- 4 nM and 0.30 +/- 0.02 microM, respectively. The IHC nAChR was both permeable to (P(Ca)/P(Na) = 8 +/- 0.9) and modulated by external Ca2+. ACh-evoked currents were potentiated by Ca2+ up to 500 microM but were reduced by higher concentrations of this cation. Ba2+ mimicked the effects of Ca2+ whereas Mg2+ only blocked these currents. In addition, elevation of extracellular Ca2+ reduced the amplitude of spontaneous synaptic currents without affecting their time course. The receptor had an EC50 for ACh of 60.7 +/- 2.8 microM in 0.5 mM Ca2+. In the absence of Ca2+, the EC50 for ACh increased, suggesting that potentiation by Ca2+ involves changes in the apparent affinity for the agonist. These pharmacological and biophysical characteristics of the IHC nAChR closely resemble those of the recombinant alpha9alpha10 nAChR, reinforcing the hypothesis that the functional nAChR at the olivocochlear efferent-IHC synapse is composed of both the alpha9 and alpha10 subunits.
A minimal biophysical model of the cochlea is used to investigate the validity of the hypothesis that a single compressive nonlinearity at the hair cell level can explain some of the suppression phenomena in cochlear responses to complex stimuli. The dependencies of the model responses on the amplitudes and frequencies of two-tone stimuli resemble in many respects the behavior of the experimental data, and can be traced to explicit biophysical parameters in the model. Most discrepancies between theory and experiment stem from simplifications in parameters of the minimal model that play no direct role in the hypothesis. The analysis and simulations predict further results which, pending experimental verification, may provide a more direct test of the influence of the compressive nonlinearity on the relative amplitudes of the synchronous response components, and hence of its role in synchrony suppression. For instance, regardless of the overall absolute levels of a two-tone stimulus applied to this type of model, the ratio of the amplitudes at the input and the ratio of the corresponding responses at the output remain approximately constant and equal (the output ratio changes by at most 6 dB in favor of the stronger tone). Other nonlinear responses to multitonal stimuli can also be reproduced, such as "spectral edge enhancement" [Horst et al., Peripheral Auditory Mechanisms (Springer, Berlin, 1985)] and some aspects of three-tone suppression [Javel et al., Mechanisms of Hearing (Monash U.P., Australia, 1983)]. In contrast to the complex behavior of suppression with increasing sound intensity and the drastic influence of the compressive nonlinearity on the absolute response measures on the auditory nerve (e.g., average rate and synchrony profiles), the percepts of complex sounds are relatively stable. This suggests that the invariant relative response measures are more likely used in the encoding and CNS extraction of the spectrum of complex stimuli such as speech.
A fundamental problem in the cellular analysis of learning and memory is the identification of the neuronal substrates of long-term information storage and their relation to short-term cellular alterations. In this report, biophysical correlates of long-term sensitization of a simple withdrawal reflex in the mollusc Aplysia were examined. A voltage-clamp analysis of the sensory neurons that control the reflex, 24 hours after sensitization training, revealed a significant reduction in net outward current. The results indicate that one mechanism for the storage of long-term sensitization is the regulation of membrane currents that influence the characteristics of the action potential and the excitability of individual neurons. The results also provide insights into the relation between short- and long-term sensitization in that the biophysical loci involved in the storage of long-term sensitization appear similar to those involved in short-term sensitization.
The metabolic response of freshly harvested, nude-mouse-derived Mycobacterium leprae to biophysical parameters was studied to facilitate an understanding of axenic culture requirements. Quantitation of intracellular ATP and the rate of [U-14C]palmitic acid incorporation into phenolic glycolipid I (PGL-I) were used as metabolic indicators after axenic incubation in modified Dubos medium under various biophysical conditions. PGL-I synthesis was optimal at 33 degrees C, whereas ATP was optimally maintained at less than or equal to 33 degrees C. Both metabolic indices showed sharp reductions at 37 degrees C. After 5 days of incubation, PGL-I synthesis and ATP maintenance showed pH optima of 5.1 to 5.6, with the higher value appearing optimal for ATP maintenance after extended incubation. Metabolic activity was negatively affected by strong reducing agents, and ATP maintenance was optimal when the gaseous environment was maintained at 2.5 to 10% oxygen. The results may partially explain the failure to cultivate the leprosy bacillus in vitro.
Biological, biochemical, and biophysical properties of three lymphocytic choriomeningitis (LCM) virus strains were compared. The biological property examined was the concentration range of virus which would, when injected into neonates, cause a carrier state. The dosage range for the CA1371 and Traub strains was found to be as broad as the limits examined (5 to 100 ld(50) units/mouse). The WCP strain, however, would only produce carriers within a 3 to 5 ld(50) range. The biochemical properties examined were the growth rates in tissue culture and the effect of varying the input ratio of virus to cells. With identical input ratios, the Traub strain reached a peak titer 32 hr after infection. The CA1371 and WCP strain reached their peaks at the 40th hr. With a 10-fold decrease in the amount of CA1371 virus per cell, peak titer (as high as in the above experiments) was not obtained until 56 hr postinfection. The biophysical properties examined were stability in density gradients and inactivation rates at 4C. In potassium tartrate gradients, full recovery of the CA1371 and WCP strain could be achieved. However, inactivation kinetics showed that only the CA1371 strain was much more stable than the Traub-LCM. The realization that marked differences in LCM strains exist is discussed in relation to certain inconsistencies in the literature.
The major internal structural protein (p30(gag)) of the Moloney leukemia virus and the endogenous Y-1 murine oncornavirus was examined for biochemical and biophysical manifestations of interstrain antigenic variation. Although the two viral proteins share murine group-specific antigenic determinants, the Y-1 virus p30 appeared to have both a lower relative number of such determinants and a decreased affinity at the cross-reactive sites for Moloney virus p30 monospecific antibodies. Further, immunological analysis indicated the presence of unique antigenic sites on the Moloney virus p30 not shared by the analogous Y-1 virus molecule. The two polypeptides copurified and had similar isoelectric points (pH 6.2 to 6.3) and sedimentation coefficients (2.47S). However, equilibrium sedimentation yielded a significant mass difference between the two proteins, 28,300 +/- 600 and 31,000 +/- 300 daltons for the Moloney and Y-1 virus molecules, respectively. Amino acid analysis indicated a concomitant increase in total residues for the Y-1 virus p30, although a number of residues appeared to have been conserved between the two viral proteins. Conformational studies and hydrodynamic calculations demonstrated marked secondary and tertiary structural differences; with the Y-1 virus p30 being an asymmetric prolate ellipsoid containing 27 to 28% alpha-helix and Moloney virus p30 being somewhat more spherical and possessing an alpha-helical content of 50 to 55%. Two-dimensional mapping of (125)I-labeled tryptic peptides of each p30 suggested that considerable sequence heterogeneity is responsible for many of the biophysical, biochemical, and immunochemical differences in these two analogous structural proteins.
Byssinosis, a lung disease that can affect cotton mill workers, may be caused in part by lipopolysaccharides (LPS) from Gram negative bacteria. In vitro, LPS complexes with sheep lung surfactant (SLS). To determine whether LPS in extracts of cotton dust alters the biophysical characteristics of lung surfactant, aqueous extracts (1.0% w:v) of sterile surgical cotton (SSC) and a bulk raw cotton dust (1182DB) were prepared. Aliquots of the soluble extracts were incubated with SLS and studied by sucrose gradient centrifugation, surface tension analysis, and high pressure liquid chromatography (HPLC). The chromatography was employed to analyse for 3-hydroxymyristate (3-HM), a fatty acid indicating LPS. Also, purified Enterobacter agglomerans LPS and 3-HM as controls and as mixtures with SLS, were studied by HPLC. Sucrose gradient centrifugation showed that SLS-SSC, SLS-1182DB, and the SLS control had similar densities that differed from the remaining controls. The SLS-1182DB exhibited a floccule absent in the other samples. Surface tension values of SLS-SSC and SLS-1182DB differed significantly from all controls but only slightly from one another. 3-Hydroxymyristate was detected by HPLC in the 3-HM control, EA-LPS, SLS-EA-LPS, and SLS-1182DB, but not in SLS-SSC or the remaining controls. Apparently, 3-HM was below the HPLC detection range in SSC. The data indicate that LPS in the 1182DB, SSC and EA-LPS samples complexed with SLS. Floccule development in SLS-1182DB but not in SLS-EA-LPS suggests a further component(s) present in the bulk raw cotton dust, as well as LPS, which complexes with SLS. The data suggest that biophysical alterations to lung surfactant may play a part in the pathogenesis of byssinosis.
We examined whether there was a threshold for change in fetal arterial PO2 to elicit alterations in plasma adrenocorticotropic hormone, arginine vasopressin, or cortisol, or to affect the incidence of fetal breathing movements or eye movements and we determined whether such a threshold changed with gestational age. Fetal sheep were exposed to two levels of hypoxemia, mild (4.6-5.3 mmHg PO2 drop) and moderate (8.3-8.8 mmHg PO2 drop) (1 mmHg = 133.322 Pa) for 1 h without pH change at 125-129 or 134-147 days of gestation within 7 days of spontaneous labor. Hypoxemia was induced by altering the inspired percent oxygen of the mother. No significant hormonal and biophysical changes were observed in mild hypoxemia at either age. In moderate hypoxemia at 125-129 days of gestation, there were significant increases of fetal adrenocorticotropic hormone, arginine vasopressin, and cortisol concentrations, and a decreased incidence of fetal breathing movements and eye movements. At 134-147 days of pregnancy, moderate hypoxemia induced a significant increase in adrenocorticotropic hormone, but the response was less than at 125-129 days of gestation. The arginine vasopressin response was similar to that at 125-129 days and there was no significant change in cortisol. There was a significant decrease in fetal breathing movements but not in eye movements. We conclude that a threshold of fetal arterial PO2 drop exists between 5 and 8 mmHg to elicit endocrine or biophysical responses to hypoxemia in fetal sheep at 125-129 days of gestation.(ABSTRACT TRUNCATED AT 250 WORDS)
Studies of Pneumocystis carinii pneumonia (PCP) suggest an important role for the surfactant system in the pathogenesis of the hypoxemic respiratory insufficiency associated with this infection. We hypothesized that PCP induces selective alterations in alveolar surfactant component expression and resultant biophysical properties. PCP was induced by intratracheal inoculation of 2 x 10(5) P. carinii organisms into C.B-17 scid/scid mice. Six weeks after inoculation, large (LA)- and small (SA)-aggregate surfactant fractions were prepared from bronchoalveolar lavage fluids and analyzed for expression of surfactant components and for biophysical activity. Total phospholipid content was significantly reduced in LA surfactant fractions from mice infected with PCP (53 +/- 15% of uninfected mice; P < 0.05). Quantitation of hydrophobic surfactant protein (SP) content demonstrated significant reductions of alveolar SP-B and SP-C protein levels in mice with PCP compared with those in uninfected mice (46 +/- 7 and 19 +/- 6%, respectively; P < 0.05 for both). The reductions in phospholipid, SP-B, and SP-C in LA fractions measured during PCP were associated with an increase in the minimum surface tension of LAs as measured by pulsating bubble surfactometer (13.1 +/- 1.1 vs. 5.4 +/- 1.8 mN/m; P < 0.05). In contrast to decreases in the hydrophobic SPs, SP-D content in the SA fraction was markedly increased (343 +/- 30% of control value; P < 0. 05) and SP-A levels in LA surfactant were maintained (93 +/- 26% of control value) during P. carinii infection. In all cases, the changes in SP content were reflected by commensurate changes in the levels of mRNA. We conclude that PCP induces selective alterations in surfactant component expression, including profound decreases in hydrophobic protein contents and resultant increases in surface tension. These changes, demonstrated in an immunologically relevant animal model, suggest that alterations in surfactant could contribute to the hypoxemic respiratory insufficiency observed in PCP.
1. Associative memory function was analyzed in a realistic biophysical simulation of rat piriform (olfactory) cortex containing 240 pyramidal cells and 58 each of two types of inhibitory interneurons. Pyramidal cell simulations incorporated six different intrinsic currents and three different synaptic currents. We investigated the hypothesis that acetylcholine sets the appropriate dynamics for learning within the network, whereas removal of cholinergic modulation sets the appropriate dynamics for recall. The associative memory function of the network was tested during recall after simulation of the cholinergic suppression of intrinsic fiber synaptic transmission and the cholinergic suppression of neuronal adaptation during learning. 2. Hebbian modification of excitatory synaptic connections between pyramidal cells during learning of patterns of afferent activity allowed the model to show the basic associative memory property of completion during recall in response to degraded versions of those patterns, as evaluated by a performance measure based on normalized dot products. 3. During learning of multiple overlapping patterns of afferent activity, recall of previously learned patterns interfered with the learning of new patterns. As more patterns were stored this interference could lead to the exponential growth of a large number of excitatory synaptic connections within the network. This runaway synaptic modification during learning led to excessive excitatory activity during recall, preventing the accurate recall of individual patterns. 4. Runaway synaptic modification of excitatory intrinsic connections could be prevented by selective suppression of synaptic transmission at these synapses during learning. This allowed effective recall of single learned afferent patterns in response to degraded versions of those patterns, without interference from other learned patterns. 5. During learning, cholinergic suppression of neuronal adaptation enhanced the activity of cortical pyramidal cells in response to afferent input, compensating for decreased activity due to suppression of intrinsic fiber synaptic transmission. This modulation of adaptation led to more rapid learning of afferent input patterns, as demonstrated by higher values of the performance measure. 6. During recall, when suppression of excitatory intrinsic synaptic transmission was removed, continued cholinergic suppression of neuronal adaptation led to the spread of excessive activity. More stable activity patterns during recall could be obtained when the cholinergic suppression of neuronal adaptation was removed at the same time as the cholinergic suppression of synaptic transmission. 7. A realistic biophysical simulation of the effects of acetylcholine on synaptic transmission and neuronal adaptation in the piriform cortex shows that these effects act together to set the appropriate dynamics for learning, whereas removal of both effects sets the appropriate dynamics for recall.
Voltage-dependent calcium channels (VDCCs) are multimeric complexes composed of a pore-forming alpha(1) subunit together with several accessory subunits, including alpha(2)delta, beta, and, in some cases, gamma subunits. A family of VDCCs known as the L-type channels are formed specifically from alpha(1S) (skeletal muscle), alpha(1C) (in heart and brain), alpha(1D) (mainly in brain, heart, and endocrine tissue), and alpha(1F) (retina). Neuroendocrine L-type currents have a significant role in the control of neurosecretion and can be inhibited by GTP-binding (G-) proteins. However, the subunit composition of the VDCCs underlying these G-protein-regulated neuroendocrine L-type currents is unknown. To investigate the biophysical and pharmacological properties and role of G-protein modulation of alpha(1D) calcium channels, we have examined calcium channel currents formed by the human neuronal L-type alpha(1D) subunit, co-expressed with alpha(2)delta-1 and beta(3a), stably expressed in a human embryonic kidney (HEK) 293 cell line, using whole cell and perforated patch-clamp techniques. The alpha(1D)-expressing cell line exhibited L-type currents with typical characteristics. The currents were high-voltage activated (peak at +20 mV in 20 mM Ba2+) and showed little inactivation in external Ba2+, while displaying rapid inactivation kinetics in external Ca2+. The L-type currents were inhibited by the 1,4 dihydropyridine (DHP) antagonists nifedipine and nicardipine and were enhanced by the DHP agonist BayK S-(-)8644. However, alpha(1D) L-type currents were not modulated by activation of a number of G-protein pathways. Activation of endogenous somatostatin receptor subtype 2 (sst2) by somatostatin-14 or activation of transiently transfected rat D2 dopamine receptors (rD2(long)) by quinpirole had no effect. Direct activation of G-proteins by the nonhydrolyzable GTP analogue, guanosine 5'-0-(3-thiotriphospate) also had no effect on the alpha(1D) currents. In contrast, in the same system, N-type currents, formed from transiently transfected alpha(1B)/alpha(2)delta-1/beta(3), showed strong G-protein-mediated inhibition. Furthermore, the I-II loop from the alpha(1D) clone, expressed as a glutathione-S-transferase (GST) fusion protein, did not bind Gbetagamma, unlike the alpha(1B) I-II loop fusion protein. These data show that the biophysical and pharmacological properties of recombinant human alpha(1D) L-type currents are similar to alpha(1C) currents, and these currents are also resistant to modulation by G(i/o)-linked G-protein-coupled receptors.
A total of 330 high-risk pregnant women with gestational ages of 32-42 weeks were followed until delivery using the biophysical profile (BPP) and a screening test consisting of the amniotic fluid index together with fetal acoustic stimulation under ultrasound M-mode scanning. The test was compared with the BPP and nonstress test (NST) for predicting abnormal outcome. The sensitivities of the BPP, NST and the proposed test were 100, 98.4 and 100%, respectively, the negative predictive values were 100, 99.4 and 100%, while the false-positive rates were 21.3, 39.3 and 67%. Our simplified biophysical testing method is suggested to be a good negative preliminary screening test, while positive results require further fetal testing methods. This test reduced the need for BPP in many high-risk patients and had the advantages of simplicity, low cost and less time consumption.
Mesangial cells, a combined smooth muscle- and fibroblast-like phenotype, are important regulators of renal function. These cells exist in a region of variable osmolarity and may require Cl(-) channels for volume regulation. Additionally, Ca(2+)-activated Cl(-) channels in these cells may participate in Ca(2+)-dependent contractile responses to vasoactive agonists. Relatively little, however, is known about mesangial cell Cl(-) currents (I(Cl)); including the biophysical description and pharmacological characterization. We used whole-cell patch clamp to study I(Cl) in cultured human and SV40-transformed murine mesangial cells. I(Cl) was measured in cells dialyzed and bathed with symmetrical N-methyl-D-glucamine chloride solutions to minimize cation currents. Dialysis with buffers to control intracellular Ca(2+) ([Ca(2+)](i)), extracellular solutions of varied osmolarity, and manipulation of the transmembrane Cl(-) gradient were used to separate two currents: I(Cl.vol) (volume-sensitive), and I(Cl.Ca) (Ca(2+)-activated). In symmetrical Cl(-) with low [Ca(2+)](i), I(Cl.vol) was outwardly rectifying and modulated by osmolarity. I(Cl.vol) demonstrated slight time- and voltage-dependent inactivation. In symmetrical Cl(-) with elevated [Ca(2+)](i) and hypertonic bath, I(Cl.Ca) was linear, but in asymmetrical Cl(-) (low [Cl(-)](i)) was outwardly rectifying and demonstrated time- and voltage-dependent activation. Permeability sequences for both I(Cl.vol) and I(Cl.Ca) were I(-) > Br(-) > Cl(-) > F(-); however, there were differences in the relative magnitudes. Tamoxifen inhibited I(Cl.vol) more potently than I(Cl.Ca), whereas niflumic acid inhibited I(Cl.Ca) more potently than I(Cl.vol). We have separated and characterized two types of I(Cl) in cultured human and murine mesangial cells. I(Cl.Ca) and I(Cl.vol) have different biophysical and pharmacological characteristics. These observations on I(Cl.Ca) and I(Cl.vol) may provide insight into mesangial cell reactivity and volume regulation.