Voltage-dependent channels permeable to K+ and Na+ in the membrane ofAcer pseudoplatanus vacuoles

Summary The patch-clamp technique in whole-cell configuration was used to study the electrical properties of the tonoplast in isolated vacuoles fromAcer pseudoplatanus cultured cells. In symmetrical KCl or K₂ malate solutions, voltage- and time-dependent inward currents were elicited by hyperpolarizing the tonoplast (inside negative), while in the positive range of potential the conductance was very small. The specific conductance of the tonoplast at –100 mV, in 100mm symmetrical KCl was about 160 S/cm². The reversal potentials (E _rev) of the current, measured in symmetrical or asymmetrical ion concentrations (cation, anion or both) were very close to the values of the K⁺ equilibrium potential. Experiments performed in symmetrical or asymmetrical NaCl indicate that Na⁺ too can flow through the channels. NeitherE _rev nor amplitude and kinetics of the current changed by replacing NaCl with KCl in the external solution. These results indicate the presence of hyperpolarization-activated channels in tonoplasts, which are permeable to K⁺ as well as to Na⁺. Anions such as Cl^– or malate seem to contribute little to the channel current.     

Tricyclic Imipramine Modification of the Circadian Rhythms of Hypothalamic Serotonin, its Precursors and Acid Catabolite in Individually Housed Rats

Circadian rhythms of serotonin (5HT), its precursors tryptophan (TP) and 5-hydroxy-tryptophan (5HTP) and its acid catabolite 5-hydroxy-indoleacetic acid (5HIAA), were determined in the hypothalamus of control rats and rats which had been treated continuously with subcutaneous imipramine (10 mg/kg/day) for 2 weeks.

Rats were individually housed and entrained to LD12:12. Controls showed the 5HT and TP peaks in the light and dark periods respectively, as reported in the literature, but no inverted correlation (antiphase) between SHT and 5HIAA rhythms.

Imipramine significantly modified circadian rhythm characteristics: the 5HT acrophase was advanced, that of TP and 5HIAA was delayed. Imipramine also significantly increased hypothalamic SHT and TP concentrations.     

Rule-based production planning in flexible manufacturing systems

Production planning in flexible manufacturing may require the solution of a large-scale discrete-event dynamic stochastic optimization problem, due to the complexity of the system to be optimized, and to the occurrence of discrete events (new orders and hard failures). The production planning problem is here approached for a multistage multipart-type manufacturing shop, where each work cell can share its processing time among the different types of parts. The solution of this problem is obtained by an open-loop-feedback control strategy, updated each time a new event occurs. At each event time, two coupled problems are solved: 1) a product-order scheduling problem, conditioned on estimated values of the production capacities of all component work cells; and 2) a production-capacity planning problem, conditioned on predefined sequences of the product orders to be processed. In particular, the article aims at defining a production planning procedure that integrates both analytical tools, derived from mathematical programming, and knowledge-based rules, coming from experience. The objective is to formulate a hybrid (knowledge-based/analytical) planning architecture, and to analyze its use for multicell multipart-type manufacturing systems.     

The β4Integrin Subunit Is Expressed in Mouse Fibroblasts and Modulated by Transforming Growth Factor-β1

Integrin β₄subunit is present in association with α₆chain on both normal and transformed epithelial cells. Recently α₆β₄heterodimer was found on the endothelium of medium-sized blood vessels and on immature thymocytes. In this report we show, by Northern blotting, indirect immunofluorescence, immunoprecipitation, and Western blotting, that β₄subunit is expressed also on cells of mesenchymal origin such as fibroblasts, myoblasts, and myotubes. Increased expression of α₆β₄has been related to the aggressive metastatic phenotype of human and murine carcinomas. The transforming growth factor β₁(TGF-β₁) has been found to modulate the expression of several integrins and intracellular matrix proteins, as well as to stimulate cell invasion and metastatic potential. To evaluate whether α₆β₄expression is modulated by TGF-β₁, we transfected 3T3 fibroblasts with an expression vector carrying the human TGF-β₁cDNA driven by the SV40 early promoter. We observed by indirect immunofluorescence a modification in the subcellular distribution of β₄subunit, which acquires a perinuclear localization. This finding suggests this integrin subunit correlates with the cytoskeletal reorganization induced by TGF-β₁.     

VAMP-2 and cellubrevin are expressed in pancreatic beta-cells and are essential for Ca(2+)-but not for GTP gamma S-induced insulin secretion.

VAMP proteins are important components of the machinery controlling docking and/or fusion of secretory vesicles with their target membrane. We investigated the expression of VAMP proteins in pancreatic beta-cells and their implication in the exocytosis of insulin. cDNA cloning revealed that VAMP-2 and cellubrevin, but not VAMP-1, are expressed in rat pancreatic islets and that their sequence is identical to that isolated from rat brain. Pancreatic beta-cells contain secretory granules that store and secrete insulin as well as synaptic-like microvesicles carrying gamma-aminobutyric acid. After subcellular fractionation on continuous sucrose gradients, VAMP-2 and cellubrevin were found to be associated with both types of secretory vesicle. The association of VAMP-2 with insulin-containing granules was confirmed by confocal microscopy of primary cultures of rat pancreatic beta-cells. Pretreatment of streptolysin-O permeabilized insulin-secreting cells with tetanus and botulinum B neurotoxins selectively cleaved VAMP-2 and cellubrevin and abolished Ca(2+)-induced insulin release (IC50 approximately 15 nM). By contrast, the pretreatment with tetanus and botulinum B neurotoxins did not prevent GTP gamma S-stimulated insulin secretion. Taken together, our results show that pancreatic beta-cells express VAMP-2 and cellubrevin and that one or both of these proteins selectively control Ca(2+)-mediated insulin secretion.