Stimulation by HCO 3 − of Na+ transport in rabbit gallbladder

Summary Bicarbonate presence in the bathing media doubles Na⁺ and fluid transepithelial transport and in parallel significantly increases Na⁺ and Cl^– intracellular concentrations and contents, decreases K⁺ cell concentration without changing its amount, and causes a large cell swelling. Na⁺ and Cl^– lumen-to-cell influxes are significantly enhanced, Na⁺ more so than Cl^–. The stimulation does not raise any immediate change in luminal membrane potential and cannot be due to a HCO ₃ ^– -ATPase in the brush border. The stimulation goes together with a large increase in a Na⁺-dependent H⁺ secretion into the lumen. All of these data suggests that HCO ₃ ^– both activates Na⁺–Cl^– cotransport and H⁺–Na⁺ countertransport at the luminal barrier.Thiocyanate inhibits Na⁺ and fluid transepithelial transport without affecting H⁺ secretion and HCO ₃ ^– -dependent Na⁺ influx. It reduces Na⁺ and Cl^– concentrations and contents, increases the same parameters for K⁺, causes a cell shrinking, and abolishes the lumen-to-cell Cl^– influx. It enters the cell and is accumulated in the cytoplasm with a process which is Na⁺-dependent and HCO ₃ ^– -activated. Thus, SCN^– is likely to compete for the Cl^– site on the cotransport carrier and to be slowly transferred by the cotransport system itself.     

The innervation of the hepatic portal vein in the rabbit: Ultrastructural evidence against “purinergic” neurotransmission

Summary The relative density of adrenergic and non-adrenergic nerves in the hepatic portal vein of the rabbit has been determined ultrastructurally. Adrenergic nerves were visualised with the modified chromaffin procedure of Tranzer and Richards (1976). Nearly equal numbers of adrenergic and non-adrenergic nerve profiles were found, indicating a much greater density of innervation by non-adrenergic nerves than that described by Burnstock et al. (1979) using light microscopic histochemical methods. These results imply that part of the argument used by Burnstock et al. (1979) to support purinergic transmission in rabbit portal vein is probably invalid.     

Direct evidence for inter-organ transport of glutathione and that the non-filtration renal mechanism for glutathione utilization involves γ-glutamyl transpeptidase

A direct examination of the inter-organ cycle of glutathione metabolism was made by determining glutathione levels in plasma obtained from various blood vessels of the rat. High levels of GSH were found in hepatic vein plasma, relative to arterial and systemic venous levels, reflecting translocation of GSH from the liver to the plasma. Renal vein plasma has a level that is 20% of arterial plasma indicating that the kidney removes glutathione from plasma not only by glomerular filtration (which can account for 20–30% of the glutathione removed), but also by a non-filtration mechanism. Inhibitors of γ-glutamyl transpeptidase decrease the fraction of glutathione removed by the kidney to a value approaching that filtered, indicating that the non-filtration mechanism involves γ-glutamyl transpeptidase.     

Factors affecting the transport of β-amino acids in rat renal brush-border membrane vesicles. The role of external chloride

The effect of a variety of ions and other solutes on the accumulation of the β-amino acid, taurine, was examined in rat renal brush-border membrane vesicles. Initial taurine uptake (15 and 30 s) is sodium-dependent with a typical overshoot. This Na⁺ effect was confirmed by exchange diffusion and gramicidin inhibition of taurine uptake. External K⁺ or Li⁺ do not increase taurine accumulation more than Na⁺-free mannitol, except that the combination of external K⁺ and Na¹ in the presence of nigericin enhances uptake. Of all anions tested, including more permeant (SCN⁻ and NO₃⁻) or less permeant (SO₄²⁻), chloride supported taurine accumulation to a significantly greater degree. Preloading vesicles with choline chloride reduced taurine uptake, suggesting that external Cl⁻ stimulates uptake. Since this choline effect could be related to volume change, due to the slow diffusion of choline into vesicles, brush-border membrane vesicles were pre-incubated with LiCl, LiNO₃ and LiSO₄. Internal LiCl, regardless of the final Na⁺ anion mixture, reduced initial rate (15 and 60 s) and peak (360 s) taurine uptake. Internal LiNO₃ or LiSO₄ with external NaCl resulted in similar or higher values of uptake at 15, 60 and 360 s, indicating a role for external Cl⁻ in taurine uptake in addition to Na⁺ effect. Although uptake by vesicles is greatest at pH 8.0 and inhibited at acidic pH values (pH less than 7.0), an externally directed H⁺ gradient does not influence uptake. Similarly, amiloride, an inhibitor of the Na⁺/H⁺ antiporter, had no influence on taurine accumulation over a wide variety of concentrations or at low Na⁺ concentrations. Taurine uptake is blocked only by other β-amino acids and in a competitive fashion. d-glucose and p-aminohippurate at high concentrations (> 10⁻³ M) reduce taurine uptake, possibly by competing for sodium ions, although gramicidin added in the presence of d-glucose inhibits taurine uptake even further. These studies more clearly define the nature of the renal β-amino acid transport system in brush-border vesicles and indicate a role for external Cl in this uptake system.     

Hydrochlorothiazide enhances the apical Cl− backflux in rabbit gallbladder epithelium: Radiochemical analysis

Hydrochlorothiazide (HCTZ) was shown to inhibit the transepithelial NaCl transport and the apical Na⁺-Cl^? symport and to depolarize the apical membrane potential in the rabbit gallbladder epithelium. The depolarization was likely related to the opening of a Cl^? conductance. To better understand whether an apical Cl^? leak is involved in the mechanism of action of HCTZ, the transapical Cl^? backflux was measured radiochemically by the washout technique. The gallbladder wall, pretreated with pronase on the serosal side to homogenize the subepithelium, was loaded with ³⁶Cl^? on the luminal side; mucosal and serosal ³⁶Cl^? effluxes (J _m , J _s ) were then measured every 2 min. The pretreatment with pronase did not alter the membrane potentials and the selectivity of the epithelium. Under control conditions and the tissue in steady-state, J _m and J _s time courses were each described by two exponential decays (A,B); the rate constants, k ^A and k ^B , were 0.71 ±0.03 and 0.16±0.01 min^?1, respectively, and correspondingly the half-times (t ₁ ^2A , t ₁ ^2B ) were 1.01±0.05 and 5.00±0.44 min (n=10); these parameters were not significantly different for J _m and J _s time courses. J _s was always greater than J _m (J _s /J _m =2.02±0.22 and 1.43 ±0.17 for A and B decays). Under SCN^? treatment in steady-state conditions, both J _m and J _s time courses were described by only one exponential decay, the component B being abolished. Moreover t ₁ ^2A was similar to that predictable for the subepithelium. It follows that it is the component B which exits the epithelial compartment. Based on the intracellular specific activity and ³⁶Cl^? J _m ^B at 0 min time of the washout experiment, the cell-lumen Cl^? backflux in steady-state was calculated to be equal to about 2 μmol cm^?2hr^?1, in agreement with the value indirectly computable by other techniques. The experimental model was well responsive to different external challenges (increases in media osmolalities; luminal treatment with nystatin). HCTZ (2.5 · 10^?4 m) largely increased ³⁶Cl^? J _m ^B . The increase was abolished by luminal treatment with 10^?4 m SITS, which not only brought back the efflux time courses to the ones observed under control conditions but even increased J _s /J _m of the cellular component, an indication of a reduced J _m ^B . It is concluded that HCTZ opens an apical, SITS-sensitive Cl^? leak, which contributes to dissipate the intracellular Cl^? accumulation and to inhibit the NaCl transepithelial transport. Moreover, the drug is likely to reduce the basal electroneutral Cl^? backflux supported by Na⁺-Cl^? cotransport, in agreement with the inhibition of the cotransport itself.     

Molecular evidence for five distinct MHC class II α genes in the rabbit

Human HLA class 11 gene probes were used to identify five distinct genes encoding the class 11 heavy chain (a chain) in the rabbit. The rabbit genes were defined by both mapping data and hybridization studies of genomic clones derived from the inbred B/J rabbit strain. Analysis of the clones by hybridization at graded stringencies indicated that one group of clones corresponded to HLA-DR, one group to HLA-DQ, and two groups to HLA-DP. Clones within a fifth group, designated DN, hybridized weakly to HLA-DR and may carry a fourth species of class II genes in the rabbit. Clones within the group showing high homology to HLA-DR were found to also contain sequences hybridizing with a probe for HLA-DR . No HLA-DP, -DQ, or -DR sequences were detected in any of the other class II clones. Distinct banding patterns observed in Southern blot analyses using either human or rabbit class II probes revealed restriction fragment length polymorphism for the different rabbit haplotypes studied. TheDN, DQ, andDR genes appear to be present as single copies whereas there are two distinctDP-like genes in the rabbit.Abbreviations used in this paper RLA major histocompatibility complex of the rabbit - RFLP restriction fragment length polymorphism - RL-5 rabbit T-cell line - SSC 0.15 M sodium chloride, 0.015 M sodium citrate     

Definitive evidence for the existence of the “sitting-atop” porphyrin complexes in nonaqueous solutions

Definitive evidence for the intermediate in metalloporphyrin formation produced in the interaction of meso-tetraphenylporphine and [Rh(CO)₂Cl]₂, the “sitting-atop” complex (SAT), is given in this article. The second-order kinetics of SAT complex formation, the small rate constant for the formation reaction, and the equilibrium study indicate a one-to-one complex between the meso-tetraphenylporphine and [Rh(CO)₂Cl]₂. The analytical data, infrared spectrum, and especially the proton magnetic resonance lead to a formulation of the “sitting-atop” complex shown     

Cellular mode of serotonin action on CI− transport in the rabbit corneal epithelium

The present work examines serotonin-induced changes in cell potential difference and barrier resistances in the corneal epithelium in vitro using voltage-measuring microelectrodes and related techniques. Component resistances were determined using voltage and resistance profiles of the epithelium before and during the serotonin response. Serotonin, added to the stromal side of the cornea in the presence of nialamide, markedly reduced transcorneal and apical membrane resistances, while basal barrier resistance increased slightly and shunt resistance was unchanged. The marked drop in apical membrane resistance after serotonin treatment reflects an increase in apical membrane chloride permeability, inasmuch as the serotonin-stimulated short-circuit current is indistinguishable from the increase in net chloride flux. Prolonged (more than 1 h) exposure of corneas to serotonin markedly depolarized the epithelial cells and reduced the voltage divider ratio from 12.3 ± 2.1 to 1.5 ± 0.5, while not significantly affecting the stimulated short-circuit current. These later effects suggest changes in epithelial ion distribution during long periods of stimulation by serotonin.     

The role of the endoplasmic reticulum in the synthesis and transport of α-amylase in barley aleurone layers

The subcellular site of -amylase (EC 1.6.2.1) synthesis and transport was studied in barley aleurone layers incubated in the presence or absence of gibberellic acid (GA₃). Using [³⁵S]methionine as a marker, the site of amino-acid incorporation into organelles isolated from aleurone layers incubated with and without GA₃ was determined following purification by isopycnic sucrose-density-gradient centrifugation. Incorporation of radioactivity into trichloroacetic-acid-insoluble proteins was greatest in those fractions exhibiting activity of an endoplasmic reticulum (ER) marker enzyme. Further fractionation of densitygradient fractions by sodium-dodecyl-sulfate polyacrylamide-gel electrophoresis showed that a major portion of the radioactivity in the ER fractions was present in a protein co-migrating with marker -amylase. This protein was identified as authentic -amylase by immunoadsorbent chromatography and affinity chromatography. The newly synthesized -amylase associated with the ER was shown to be sequenstered within the lumen of the ER by experiments which showed that the enzyme was resistant to proteolytic degradation. The labelled -amylase sequestered in the ER can be chased from this organelle when tissue is incubated in unlabelled methionine following a 1-h pulse of labelled methionine. The isoenzymic forms of -amylase found in tissue homogenates and incubation media of aleurone layers incubated with and without GA₃ were characterized after chromatography on diethylaminoethyl cellulose. In homogenates of GA₃-treated aleurone layers, five peaks of -amylase activity were detected, while in homogenates of aleurone layers incubated with-out GA₃ only three peaks of activity were found. In incubation media, four isoenzymes were found after GA₃ treatment and two were found after incubation without GA₃. We conclude that at least five -amylase isoenzymes are synthesized by the ER of barley aleurone layers and that this membrane system is involved in the sequestration and transport of four of these isoenzymes.Abbreviations CHA cyclohepataamylose - DEAE-cellulose diethylaminoethyl-cellulose - ER endoplasmic reticulum - GA₃ gibberellic acid - SDS-PAGE sodium-dodecyl-sulfate polyacrylamide-gel electrophoresis     

Regulation and molecular mechanisms of calcium transport genes: do they play a role in calcium transport in the uterine endometrium?

Maintenance of calcium (Ca) balance in the uterus is critically important for many physiological functions, including smooth muscle contraction during embryo implantation. Ca transport genes, i.e., transient receptor potential cation channel subfamily V members 5/6 (TRPV5/6), calbindins, plasma membrane Ca(2+)-ATPase 1 (PMCA1), and NCX1/NCKX3, may play roles in the uterus for Ca transport and reproductive function. Although these Ca transport genes may have a role in Ca metabolism, their role(s) and molecular mechanisms require further elucidation. In this review, we highlight the expression and regulation of Ca transport genes in the uterus to clarify their potential role(s). Since Ca transport genes are abundantly expressed in reproductive tissues in a distinct manner, they may be involved in specific uterine functions including fetal implantation, Ca homeostasis, and endometrial cell production.     

Immunocytochemical evidence for the presence of “mammalian” neurohormonal peptides in neurones of the tapeworm Diphyllobothrium dendriticum

Summary In the nervous system of the obligatory endoparasite Diphyllobothrium dendriticum immunoreactivity (IR) to growth hormone-releasing factor (GRF), peptide histidine isoleucine (PHI), bovine pancreatic polypeptide (BPP), gastrin, gastrin-releasing peptide (GRP), oxytocin, FMRF-amide (FMRF) and serotonin (5HT) was demonstrated by immunocytochemical methods. A very strong GRF-IR was observed in the CNS and PNS of larvae and of the constantly growing adult worms. GRF-IR axon terminals occur beneath the basal lamina of the tegument along the inside of the bothridia, the holdfast organ of the worm. GRF-IR fibres surround the yolk producing vitelline glands and occur in the wall of the vagina. PHI-IR was observed in the CNS and PNS of larvae and adult worms. PHI-IR terminals occur beneath the basal lamina of the tegument along the strobila, the nutrient absorbing surface of the worm. PHI-IR fibres seem to innervate the testicular follicles. FMRF-IR fibres and perikarya occur close to the vitelline glands and the uterine pore and in the male copulatory organ. Numerous large 5HT-IR perikarya with long varicose fibres were observed in the nervous system of the worm. 5HT-IR perikarya occur close to the genital atrium. D. dendriticum is the phylogenetically lowest organism in which IR to PHI has been demonstrated.     

Detritivores in Kenyan highland streams: more evidence for the paucity of shredders in the tropics?

1. The relationship between coarse particulate organic matter (CPOM) standing stock and benthic macroinvertebrate assemblages in Kenyan highland streams was determined by sampling seven sites on three rivers (2000–2700 m a.s.l.). Taxa recorded were allocated to functional feeding groups using published literature, mouthpart analysis and examination of gut contents. Patterns were compared with five structurally similar streams in three areas of Europe (south-west France, south-east England, north-east England).
2. Number of individuals and proportion of detritivores in Kenyan streams were equivalent to, or greater than, those in European sites. Shredders were, however, almost completely absent from Kenyan sites, despite high standing stocks of CPOM. Shredders were abundant in all European sites.
3. The phenomenon of low shredder abundance has been observed in other tropical streams in south-east Asia and Central and South America but, in contrast to these regions, the African rivers studied were devoid of shrimps or fish which may occupy the shredding niche elsewhere.
4. These preliminary data suggest that shredder-mediated detritus processing, which is a key functional component of streams in the North Temperate Zone, does not operate in East African streams. There are three possible reasons for this. The first is that tropical African rivers are functionally different to those in temperate regions. This could be because of enhanced microbial activity replacing shredder activity at high temperatures. Alternatively, it could be a result of low palatability of detrital inputs from dominant riparian trees in the region. The second and third are methodological: that our allocation to functional feeding groups is incorrect, and that our sampling methods missed a potentially key shredding taxon – the freshwater crab Potamonautes sp.     

Intracellular localization of inositol-phospholipid-metabolizing enzymes in rabbit fast-twitch skeletal muscle. Can D-myo-inositol 1,4,5-trisphosphate play a role in excitation-contraction coupling?

Rabbit fast-twitch skeletal muscle microsomes have been separated by isopycnic centrifugation on a linear sucrose gradient into triads and light sarcoplasmic reticulum. In both fractions phosphatidylinositol-kinase activity is found [Varsányi et al. (1986) Biochem. Biophys. Res. Commun. 138, 1395]. In contrast, phosphatidylinositol-4-phosphate kinase is nearly exclusively associated with triads. The phosphatidylinositol-4,5-bisphosphate-phosphodiesterase activity shows a biphasic distribution: approximately 50% of the activity is associated with triads and 50% appears in the overlay. Triads have been broken mechanically into transverse tubules and terminal cisternae, then separated by isopycnic sucrose-gradient centrifugation. Both fractions exhibit phosphatidylinositol-kinase activity; the activities of phosphatidylinositol-4-phosphate kinase and phosphatidylinositol-4,5-bisphosphate phosphodiesterase are associated mainly with the transverse tubules. Consequently, in rabbit fast-twitch skeletal muscle all necessary enzymes for production of D-myo-inositol 1,4,5-trisphosphate are associated with transverse tubules. Phosphatidylinositol-4,5-bisphosphate phosphodiesterase associated with triads shows a pH optimum at 6.8. The enzyme is maximally active between pCa 5 and pCa 4. Mg2+ inhibits the enzyme activity half-maximally at about 1 mM. Guanine-nucleotide-binding proteins seem not to be involved in the regulation of enzyme activity; guanosine 5'-[gamma-thio]triphosphate does not influence phosphatidylinositol-4,5-bisphosphate phosphodiesterase activity. It correlates well with the observation that neither alpha 1-adrenergic nor muscarinic receptors have been found in fast-twitch rabbit skeletal muscle. On basis of the respective enzyme activities estimations on maximal phosphatidylinositol turnover were made and a possible involvement of this signal pathway in excitation-contraction coupling has been discussed. Furthermore, calculations show that during a single twitch D-myo-inositol 1,4,5-trisphosphate concentration does not reach more than 2 nM. However, during a 4-s tetanus D-myo-inositol 1,4,5-trisphosphate can accumulate to a level which could effect force generation [Thieleczek and Heilmeyer (1986) Biochem. Biophys. Res. Commun. 135, 662] and aldolase distribution (Thieleczek et al., unpublished results).     

Electroencephalographic evidence of “Gestalt” in the perception of movement by the frog

Summary Some records, obtained from the surface of the optic tectum of the frog with moving visual stimuli are presented as evidence of a global oscillation of the tectal activity whose time course is specific for different patterns of stimulation.The research reported in this document has been sponsored by the 6570th Aerospace Medical Research Laboratory under grant AF EOAR 65-44 through the European Office of Aerospace Research (OAR), United States Air Force.     

A model for the perception of curves in dot figures: The role of local salience of “virtual lines”

In many models of visual information processing the notion of a virtual line or dipole is introduced in order to represent the configurational information, notably length and relative orientation, between identical figure elements in figures with discrete elements. Virtual lines have proven to be very useful in predicting perceptual phenomena (Julesz et al. 1973; Stevens 1978). In the present study, virtual lines are utilized in a model which aims to predict the perception of (dotted) curves in dot figures. Clearly many possible curves, formed by adjacent virtual lines, can be constructed within a set of dots. It is proposed that already at the local level of the virtual lines each line has a perceptual salience which results from the function induced by the global dot figure. It is this local line salience or connectivity that directs further processing and determines the curves to be seen in a dot figure. The model presented is an information processing model with a clear modular design. It entails three successive levels of representation. First image functions are derived through a convolution of the input with gaussian distribution functions. Next, a discrete internal representation is extracted from the image function consisting of two primitives; blobs, representing the dots, and virtual lines, representing pairwise relations between blobs. The attributes of the blobs are their positions in the image plane, while those of the virtual lines are length, relative orientation and connectivity. At the third level, the discrete internal representation is used to predict the perceived curves. It is shown that the model has advantages over other approaches, e.g. autocorrelation and network models.