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M Joffre

Publications and source records attributed to M Joffre.

At least 37 records · Page 2Linked to original sources
PubMed1996

Influence of the molecular structure of steroids on their ability to interrupt gap junctional communication.

17 beta-estradiol propionate was found to reduce the gap junctional communication in a concentration range similar to that of testosterone propionate, in primary cultures of rat Sertoli cells and cardiac myocytes. Uncoupling was reversible on washing out and occurred without concomitant rise in the intracellular calcium concentration. Esterification was prerequisite for the activity of extracellularly applied steroid compounds (for example, testosterone was ineffective even at external concentrations up to 100 microM, whereas its intracellular application at 1 microM totally interrupted intercellular communication), but their uncoupling efficiency did not depend on the nature of the ester chain nor on its position on the steroid nucleus. The derivatives of two other androgen hormones (derivatives of the androstane nucleus) were also efficient as junctional uncouplers. Among five steroid molecules belonging to the pregnane family, only one (pregnanediol diacetate) interrupted the junctional communication. Neither cholic acid nor cholesteryl acetate or ouabain showed this effect. Altogether, no correlation with the presence or position of double bonds nor with the trans- or cis-fusion of the A and B rings could be recognized. These results suggest that this reversible, nondeleterious uncoupling effect of steroids is independent of the shape of the molecules and is more probably related to their size and liposolubility, that condition their insertion into the lipid bilayer. Their incorporation into the membrane could disturb the activity of the membrane proteins by a physical mechanism.

Calcium↗
PubMed1996

Swelling and cAMP on hyperpolarization-activated Cl- conductance in rat Leydig cells.

We have used the whole cell patch-clamp technique to characterize changes in membrane conductance induced by osmotic swelling in mature rat Leydig cells dialyzed with ATP (control cells) or adenosine 3',5'-cyclic monophosphate (cAMP) plus ATP (cAMP cells). A spontaneous current activation occurs in both groups in isosmotic conditions (300/295 mosM in/out). This development is entirely counteracted in control cells and partly inhibited in cAMP cells by exposing them to a hyperosmotic (350 mosM) bath solution, and these currents increase again in a hyposmotic (205 mosM) bath solution. These currents are sensitive to 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid, a Cl- channel blocker. Taken together, the results indicate that, in the control cells (ATP alone) as well as in the presence of intracellular cAMP, osmotic swelling activates the background hyperpolarization-activated Cl- conductance, osmotic swelling and cAMP appearing to act synergistically.

Animals↗
PubMed1994

Follicle-stimulating hormone increases gap junction communication in Sertoli cells from immature rat testis in primary culture.

The gap junction communication in Sertoli cells from immature rat testes, cultured either in absence or in presence of follicle-stimulating hormone (FSH), was studied by microinjection of a fluorescent dye and by Fluorescence Recovery After Photobleaching (gapFRAP). The cells cultured for 2-4 days in the absence of FSH showed a flattened "epithelial-like" appearance. They were poorly coupled, as judged by the low frequency of cell-to-cell spread of microinjected Lucifer Yellow, and by the value of the rate constant of dye transfer (k) estimated in gapFRAP experiments. However, when two different subpopulations of cells were separately analyzed, namely the cells forming small groups contacting over part of their circumference ("adjoining cells"), and the cells arranged in tight clusters, we found that the value of k in the latter group was much higher, reaching about 75% of that obtained in the presence of FSH. The cells cultured for two days in a medium containing ovine FSH underwent striking morphological changes and presented a rounded, "fibroblast-like" appearance. They were arranged in networks or in clusters. The frequency of cell-to-cell dye diffusion after microinjection and the rate constant of dye transfer were rapidly increased to the same final level by FSH, although they were initially different in these two groups. A concentration dependence of k, in the range 0.05 to 3 ng/ml, was observed in the cells in networks, contrasting with an all-or-none increase in the cells in clusters. Two days after FSH withdrawal, the dye transfer constant returned to prestimulation control values in the cells in clusters, but not in the cells in networks, which maintained a stable degree of coupling comparable to that of the unstimulated cells in clusters. This observation suggests (i) that an initial promoting effect of FSH already exists in the immature rat testis, which is preserved after enzymatic treatment in the cell clusters, but not in the more dispersed cells, and (ii) that the decreased junctional coupling is re-established in the dispersed cells by FSH, through a synthesis or a membrane insertion of connexin. The effects of FSH were mimicked by a brief exposure to 1 mM dibutyryl-cyclicAMP, but not to 10 nM human chorionic gonadotropin (hCG), indicating that the gap junction communication in Sertoli cells is upregulated by FSH through a specific membrane receptor, with cyclicAMP acting as a second messenger.

Animals↗
PubMed1993

Characterization and cyclic AMP-dependence of a hyperpolarization-activated chloride conductance in Leydig cells from mature rat testis.

We recently described a cyclic AMP-activated current in the membrane of Leydig cells from mature rat testis by using the whole-cell configuration of the patch-clamp technique (Noulin & Joffre, 1992a). In the present study, further experiments were performed in symmetrical CsCl solutions. We show that this current corresponds to a hyperpolarization-activated chloride conductance. Voltage jumps to negative potentials, applied from a holding potential of +60 mV, activated a time-dependent inward current. In control cells, the curve of steady-state current activation typically ranged from +60 mV (0) to -120 mV (1) and had its midpoint at -40 mV. Deactivation at positive potential was characterized by an instantaneous outwardly rectifying current which decayed with time. The kinetics of activation and deactivation were described by a double and a single exponential, respectively. Cyclic AMP, added to the pipette solution, increased both the inward rectification and the amplitude of the steady-state current in the range of 0 to -60 mV. The activation threshold was unchanged, while the V0.5 of the activation curve was shifted by 24 mV to more positive potentials. Consequently, the activation curve was steeper. The two rate constants of activation were increased and were strongly voltage dependent. In parallel, the amplitude of the instantaneous outward current and the rate constant of deactivation were increased. The reversal potential of this current was close to ECl. It did not change with equimolar replacement of cesium by TEA, and shifted with the chloride concentration gradient. This current was inhibited by chloride channel blockers. These results indicate a hyperpolarization-activated chloride conductance in the membrane of Leydig cells which is modulated by cyclic AMP. This nucleotide acts by modifying the kinetics of inward current and both the kinetics and the amplitude of deactivating outward current.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1991

[Increase by FSH hormone and depression by testosterone of the diffusional coupling between Sertoli cells from immature rat testis in primary culture].

Previous studies have suggested that FSH promotes the intercellular coupling of Sertoli cells from immature rat testis in primary culture ([1], [2]). In order to test this hypothesis, we have investigated the diffusional coupling between Sertoli cells in primary culture with the FRAP technique. The coupling is low in unstimulated cells but increases in the presence of FSH. This effect is not reversed by returning to the control medium. Testosterone decreases this coupling, an effect which is reversed by a new exposure to FSH. Taken together these data show that FSH initiates diffusional coupling in Sertoli cells and that testosterone antagonizes this effect.

Animals↗
PubMed1990

Glucose- and concentration-dependence of noradrenalin effects on electrical activity in mouse pancreatic beta cells.

The effects of a wide range of noradrenalin (NA) concentrations (10(-11)-10(-4) M) on the membrane potential and on the glucose-induced electrical activity were investigated with microelectrodes in microdissected mouse islets. In the presence of 11.1 mM glucose, the beta cells exhibited a repetitive activity. NA at more than 10(-7) M induced a rapid hyperpolarization followed by a silent depolarization, then by the appearance of a slowed pace of repetitive activity (dose-dependent effects). NA at less than 10(-7) M did not markedly affect the electrical activity; it only induced a dose-dependent increase in the degree of activity with no change of the potential levels. The glucose-dependence of these effects were then investigated. In the absence of glucose, 10(-8) and 10(-6) M NA did not affect the resting membrane potential. Non-stimulatory glucose concentrations (2.8-7.3 mM) progressively decreased the membrane potential. 10(-8) M NA did not affect it, while 10(-6) M NA induced a dose-dependent and long-lasting hyperpolarization. In the presence of stimulatory glucose concentrations (7.3-30 mM) the degree of activity increased, 10(-8) M NA induced a slight leftward shift and 10(-6) M NA a slight rightward shift of the dose-response curve.

Animals↗
PubMed1990

Potassium and chloride conductances in rat Leydig cells: effects of gonadotrophins and cyclic adenosine monophosphate.

1. The effects of gonadotrophins (luteinizing hormone and human chorionic gonadotrophin) and cyclic AMP on ionic conductances were investigated using the tight-seal whole-cell recording technique in Leydig cells freshly isolated from nature rat testis by enzymatic treatment. 2. In resting cells, the predominant ionic conductance is a voltage-dependent K+ conductance resembling the delayed rectifier K+ conductance of T-lymphocytes. This conductance is characterized by: (1) a time-dependent inactivation for potentials more positive than +20 mV, (2) a reversal potential near -65 mV, (3) a sensitivity to intracellular Cs+, and (4) a sensitivity to extracellular TEA and 4-aminopyridine. 3. A Cl- conductance is also present resembling the Cl- background conductance in squid axons and heart cells. In resting cells, this conductance contributes only a small component of the total outward current obtained with depolarizing pulses. 4. Gonadotrophins (human chorionic gonadotrophin, porcine luteinizing hormone and ovine luteinizing hormone) have little effect on the K+ conductance. They transiently increase a Cl- conductance after a delay of up to 30 s. This response does not occur if the hormones are applied late in the whole-cell recording. Gonadoliberine (GnRH) does not affect the Cl- or K+ conductance. 5. Internal cyclic AMP (100 microM) mimics all these effects while internal application of a GTP-ATP mixture induces a similar response, which is, however, sustained rather than transient. 6. The Cl- conductance was studied quantitatively with a GTP-ATP internal solution. This conductance is activated by depolarizing voltage steps to test potentials of -40 mV or more. Under these conditions, the instantaneous current observed as soon as the depolarizing pulse is applied displays outward rectification and reverses near ECl. During the pulses, a strong inactivation is observed for potentials greater than +40 mV. This conductance is independent of external and internal calcium. 7. It is concluded that the gonadotrophins act through a cyclic AMP-dependent process to activate a Cl- conductance. This conductance is different to the hyperpolarization-activated Cl- conductance and the calcium-activated Cl-conductance also present in the membrane of resting cells.

Animals↗
PubMed1989

Effect of uncoupling treatments on FSH-induced hyperpolarization of immature rat Sertoli cells from Sertoli cell-enriched cultures.

Sertoli cell-enriched cultures isolated from immature rat testes by enzymic treatments were investigated by intracellular microelectrode recordings. The hyperpolarization of cells induced by FSH was independent of the age of the rats (7-37 days) and was unchanged by exposure to a hormone-free medium or to a glycine buffer of pH 3. It was reduced by treatments which decreased the electrical coupling between cells either by an increase of intracellular calcium [i.e. calcium ionophore (A 23187, 5 x 10(-6) M), general anaesthetic (heptanol, 3.5 mM) and uncoupler of oxidative phosphorylations (carbonylcyanide m-chlorophenylhydrazone-CCmP, 10(-6) M)] or by a decrease of extracellular calcium [i.e. 0Ca + EGTA (1 mM) medium]. These effects were partly or totally reversed by a recovery period in a drug-free medium. Similar results were obtained by an exposure to trypsin (0.05%) followed by a second mechanical dispersion, but new cell hyperpolarization was induced by a new exposure to FSH. This electrophysiological study suggests an initial effect of FSH on the junctional complex between Sertoli cells, then the control by calcium of this complex.

Aging↗
PubMed1988

Electrophysiological studies of the action of gonadotropins on Leydig and Sertoli cells from rat testis.

Using intracellular microelectrode recordings, it was observed that FSH caused the hyperpolarization of cultured Sertoli cells from the testis of 17-day old rats; this hyperpolarization was related to the activation of a Na/K pump and an increase in Na/Ca influx and K efflux. The effect was irreversible and mediated by cAMP. Some other results suggested an initial effect of FSH on the junctional complex between the Sertoli cells and then its calcium dependence. Voltage-clamped membrane currents were investigated from whole-cell patch clamp recordings performed on single Leydig cells isolated from adult rat testis. Two outward currents were evoked by depolarizing voltage steps: a potassium current recorded in cells dialyzed with low-calcium media (10(-9)-10(-8) M) and a chloride current recorded in cells dialyzed with high-calcium media (10(-7)-10(-6) M). These responses to modifications of intracellular calcium were not affected by hCG. However, this hormone induced an early change of the K current to a Cl current when the cells were dialyzed by a low-buffered EGTA-calcium medium. These results suggest that hCG stimulation involved the blockade of a potassium current and the activation of a chloride current through an increase of intracellular calcium.

Animals↗
PubMed1988

Follicle-stimulating hormone induces hyperpolarization of immature rat Sertoli cells in monolayer culture.

1. The effect of FSH on the membrane potential of Sertoli cells has been investigated by intracellular microelectrode recording from monolayer cultures of cells, isolated from immature rat testes by enzymatic treatment. 2. In standard Earle's solution, the membrane potential of unstimulated cells in a 3-day-old monolayer was -21.6 +/- 0.2 mV (n = 300). The recorded potentials were unchanged on varying the culture period from 3 to 7 days or by removal of chloride or calcium from the media. However, they were decreased in a low-sodium or potassium-rich medium. 3. Ovine FSH caused hyperpolarization of the cell in a dose-dependent manner. Maximal values were obtained with concentrations ranging between 2.9 and 5.9 micrograms/ml (-37.0 +/- 0.2 mV; n = 310). Dibutyryl cyclic AMP (1 mM) produced a similar effect to FSH. Under similar conditions human chorionic gonadotrophin (hCG) had no effect on the membrane potential. 4. The FSH-induced hyperpolarization was unchanged by chloride or bicarbonate replacement. It was decreased by low-sodium media (9 mM) (-29.6 +/- 0.3 mV; n = 110), by removal of calcium (-32.4 +/- 0.6 mV; n = 128) and by an increase in the potassium concentration of the bathing medium. A tenfold increase in potassium concentration depolarized the cell by 29.5 mV against 16 mV for unstimulated cells. 5. FSH-induced hyperpolarization was decreased by application of ouabain (10(-4) M), quinidine (1.4 x 10(-4) M) and cobalt (10(-3) M) (respectively: -28.7 +/- 0.3 mV, n = 124; -26.6 +/- 0.3 mV, n = 52 and -24.8 +/- 0.3 mV, n = 68) but was unchanged by amiloride (10(-4) M) and TTX (3 x 10(-6) M). None of these drugs modified the membrane potential of unstimulated cells. 6. All these results suggest that FSH stimulation of Sertoli cells in monolayer culture involves the modification of their membrane permeabilities.

Animals↗
PubMed1987

Ca-dependent-chloride and potassium currents in rat Leydig cells.

Voltage-clamped membrane currents have been investigated from whole-cell patch-clamp recordings performed on single Leydig cells isolated from the adult rat testis. Two outward membrane currents were evoked by depolarizing voltage steps. A potassium current was recorded in cells dialyzed with low (10(-9)-10(-8) M) calcium media. This current was decreased by TEA (10 mM). A chloride current was recorded in cells dialyzed with high (10(-7)-10(-6) M) calcium media. This current was decreased by an external exposure to glutamate. Comparison of the currents at low and high internal calcium concentrations suggests that an increase of the intracellular calcium activates a chloride current.

Animals↗
PubMed1987

Sensitivity of pancreatic beta cell to calcium channel blockers. An electrophysiologic study of verapamil and nifedipine.

Microelectrodes were used to study the comparative effects of 2 calcium channel blockers on glucose-induced electrical activity in mouse beta cells. In 2.8 mM glucose, verapamil (10(-5) M), but not nifedipine (10(-7) M), induces a silent depolarization. In 11.1 mM glucose, verapamil (10(-7) to 5.10(-5) M) induces continuous spike activity by a decrease in the maximum repolarization potential. Nifedipine (10(-10) to 10(-6) M) induces the same activity, but subsequent to a hyperpolarization of the cell at the maximal repolarization potential followed by a silent phase to the plateau potential. The 2 drugs induce a dose-dependent decrease in spike frequency without any change in spike amplitude. In 22 mM glucose exposure to nifedipine, but not to verapamil, induces a transient period of slow-wave activity. The 2 drugs induce a dose-dependent decrease in spike frequency. At higher concentrations (nifedipine greater than 10(-7) M; verapamil greater than 10(-6) M) they induce the disappearance of spikes through a decrease in amplitude. These results show that the beta cell is more sensitive to nifedipine (ED50 = 3 X 10(-8) M) than to verapamil, and that glucose stimulation increases the cell's sensitivity to verapamil (11.1 mM glucose: ED50 = 10(-5) M versus 5 X 10(-7) M in 22 mM glucose) but not to nifedipine.

Animals↗