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J M Israel

Publications and source records attributed to J M Israel.

At least 19 recordsLinked to original sources
PubMed2000

17-Oestradiol modulates in vitro electrical properties and responses to kainate of oxytocin neurones in lactating rats.

1. Intracellular current clamp recordings were performed from identified oxytocin (OT) neurones in acute hypothalamic slices taken from lactating Wistar rats at early (5th day: LD-5) and late (21st day: LD-21) lactation. 2. The basic electrophysiological properties of LD-21 OT neurones differed from those of LD-5 OT neurones: their resting membrane potential was more depolarised (-51.5 versus -54.9 mV); their action potential duration was longer (1.6 versus 1.2 ms); their hyperpolarising after-potential (HAP) following single spikes and after-hyperpolarisation (AHP) following a burst of action potentials had smaller amplitudes (-46 and -67 %, respectively); and they lacked spike frequency adaptation during a burst. 3. In LD-21 neurones bath application of 17beta-oestradiol (10-7 M, 6-14 min) reversibly restored all these properties to values observed in LD-5 cells. This treatment had no effect on LD-5 neurones. 4. LD-21 neurones were less sensitive to kainate than LD-5 neurones. 17beta-Oestradiol significantly potentiated the kainate-induced response in LD-21, but not in LD-5 neurones. 5. The effects of 17beta-oestradiol were presumably mediated through a non-genomic mechanism since they occurred within a few minutes of administration, and disappeared within 30-40 min of washout. They were not inhibited by tamoxifen, an antagonist of the nuclear oestrogen receptor ER-alpha. Lastly, cholesterol, a non-active lipophilic molecule, had no effect. 6. Our observations demonstrate that, in the absence of 17beta-oestradiol, the basic electrical properties and sensitivity to kainate of OT neurones become altered between early and late lactation. However, the rise in circulating levels of oestrogens during the late phase of lactation may contribute to maintain OT neurone reactivity as long as suckling continues.

Animals↗
PubMed2000

Spontaneous neuronal activity in organotypic cultures of mouse dorsal root ganglion leads to upregulation of calcium channel expression on remote Schwann cells.

It is well established that neurons regulate the properties of both central and peripheral glial cells. Some of these neuro-glial interactions are modulated by the pattern of neuronal electrical activity. In the present work, we asked whether blocking the electrical activity of dorsal root ganglion (DRG) neurons in vitro by a chronic treatment with tetrodotoxin (TTX) would modulate the expression of the T-type Ca(2+) channel by mouse Schwann cells. When recorded in their culture medium, about one-half of the DRG neurons spontaneously fired action potentials (APs). Treatment for 4 days with 1 microM TTX abolished both spontaneous and evoked APs in DRG neurons and in parallel significantly reduced the percentage of Schwann cells expressing Ca(2+) channel currents. On the fraction of Schwann cells still expressing Ca(2+) channel currents, these currents had electrophysiological parameters (mean amplitude, mean inactivation time constant, steady-state inactivation curve) similar to those of control cultures. Co-treatment for 4 days with 1 microM TTX and 2 mM CPT-cAMP, a cAMP analogue that induces the expression de novo of Ca(2+) channel currents in Schwann cells deprived of neurons, maintained the percentage of Schwann cells expressing Ca(2+) channel currents, showing that TTX does not directly affect the expression of Ca(2+) channel currents by Schwann cell. We conclude that blocking spontaneous activity of DRG neurons in vitro downregulates Ca(2+) channel expression by Schwann cells. These results strongly suggest that DRG neurons upregulate Ca(2+) channel expression by Schwann cells via the release of a diffusible factor whose secretion is dependent on electrical activity.

Animals↗
PubMed1999

Visualization of local afferent inputs to magnocellular oxytocin neurons in vitro.

We recently showed that oxytocin (OT) neurons in organotypic slice cultures obtained from postnatal rat hypothalamus display complex patterns of electrical activity, similar to those of adult magnocellular OT neurons in vivo. Here we used such cultures to investigate the identity and, in particular, the origin of afferent inputs responsible for this activity. Multiple immunostaining with light and confocal microscopy showed that the somata and dendrites of oxytocinergic neurons were contacted by numerous synapses, visualized by their reaction to the synaptic markers, synaptophysin or synapsin. Many were GABAergic, displaying immunoreactivities for glutamic acid decarboxylase or gamma-aminobutyric acid (GABA); others were enriched in glutamate immunoreactivity. Such afferents presumably arose from GABA- or glutamate-immunoreactive neurons, respectively, with distinct and characteristic morphologies and topographies. A few dopaminergic boutons (tyrosine hydroxylase- or dopamine-immunopositive) impinged on OT neurons; they arose from dopamine-positive neurons located along the third ventricle. No noradrenergic profiles were detected. Despite the presence of choline acetyl-transferase (ChAT)-immunoreactive neurons, there were no cholinergic contacts. Lastly, we found oxytocinergic synapses, identified by immunoreaction for OT-related neurophysin and synapsin, contacting OT somata and dendrites. Our observations thus demonstrate that inhibitory and excitatory inputs to OT neurons derive from local intrahypothalamic GABA and glutamate neurons, in close proximity to the neurons. They also reveal that OT neurons are innervated by hypothalamic dopaminergic neurons. Finally, they confirm the existence of homotypic OT synaptic contacts which derive from local OT neurons.

Acetylcholinesterase↗
PubMed1998

Evidence for a hypothalamic oxytocin-sensitive pattern-generating network governing oxytocin neurons in vitro.

During lactation and parturition, magnocellular oxytocin (OT) neurons display a characteristic bursting electrical activity responsible for pulsatile OT release. We investigated this activity using hypothalamic organotypic slice cultures enriched in magnocellular OT neurons. As shown here, the neurons are functional and actively secrete amidated OT into the cultures. Intracellular recordings were made from 23 spontaneously bursting and 28 slow irregular neurons, all identified as oxytocinergic with biocytin and immunocytochemistry. The bursting electrical activity was similar to that described in vivo and was characterized by bursts of action potentials (20.1 +/- 4.3 Hz) lasting approximately 6 sec, over an irregular background activity. OT (0.1-1 microM), added to the medium, increased burst frequency, reducing interburst intervals by 70%. The peptide also triggered bursting in 27% of nonbursting neurons. These effects were mimicked by the oxytocin receptor (OTR) agonist [Thr4, Gly7]-OT and inhibited by the OTR antagonist desGly-NH2d(CH2)5[D-Tyr2,Thr4]OVT. Burst rhythmicity was independent of membrane potential. Hyperpolarization of the cells unmasked volleys of afferent EPSPs underlying the bursts, which were blocked by CNQX, an AMPA/kainate receptor antagonist. Our results reveal that OT neurons are part of a hypothalamic rhythmic network in which a glutamatergic input governs burst generation. OT neurons, in turn, exert a positive feedback on their afferent drive through the release of OT.

Animals↗
PubMed1998

Electrophysiological studies of oxytocin neurons in organotypic slice cultures.

We have developed organotypic slice cultures derived from postnatal rat hypothalamus which contain well-differentiated oxytocin neurons. Intracellular recordings of identified neurons show that these cultured oxytocin cells exhibit basal electrical properties closely similar to those of magnocellular cells recorded in vivo and in acute in vitro preparations from adult animals. The cultures also include GABAergic and glutamatergic neurons making connections with the oxytocin cells, which strongly suggests that the rich GABAergic and glutamatergic innervations of adult oxytocin neurons in vivo derive largely from local hypothalamic sources. Pharmacological manipulations indicate that the cultured oxytocin neurons present functional GABAA (but not GABAB) receptors, and ionotropic non-NMDA and NMDA receptors, but no metabotropic receptors for glutamate. These synaptic inputs control to a great extent the electrical activity of oxytocin neurons. Of particular interest is our observation that the cultured oxytocin neurons display a recurrent bursting activity which does not appear to result from an endogenous regenerative activity, but from a patterned glutamatergic input. Our preliminary data show that oxytocin plays a facilitatory role in this bursting activity and suggest that such activity is generated within an hypothalamic circuitry.

Animals↗
PubMed1996

Electrical properties of oxytocin neurons in organotypic cultures from postnatal rat hypothalamus.

1. Intracellular recordings were performed on immunocytochemically identified oxytocin (OT) neurons (n = 101) maintained for 2-7 wk in hypothalamic organotypic cultures derived from 4-to 6-day-old rat neonates. The neurons displayed a resting potential of -58.9 +/- 6.8 mV (mean +/- SD, n = 74), an input resistance of 114 +/- 26.8 M omega (n = 66), and a time constant of 9.6 +/- 1.4 ms (n = 57). Voltage-current (V-I) relations, linear at resting potential, showed a pronounced outward rectification when depolarized from hyperpolarized membrane potentials. At these hyperpolarized potentials, depolarizing current pulses induced a delayed action potential. 2. Action potentials had an amplitude of 73.4 +/- 9.7 mV and a duration of 1.9 +/- 0.2 ms. Each action potential was followed by an afterhyperpolarization of 7.9 +/- 2.0 mV in amplitude lasting 61.7 +/- 11.3 ms. The depolarizing phase of action potentials was both Na+ and Ca2+ dependent, whereas repolarization was due to a K+ conductance increase. 3. When Ba2+ was substituted for Ca2+ in the medium, OT neurons displayed prolonged sustained depolarizations. In the presence of tetrodotoxin (TTX), these depolarizations were triggered by depolarizing current pulses and arrested by hyperpolarizing current pulses or by local application of Ca2+, Co2+, Cd2+, No sustained depolarization was obtained when nifedipine was added to the medium. These data suggest that OT cells in organotypic culture possess L-type Ca2+ channels. 4. All OT neurons generated spontaneous action potentials at resting potential. Of 59 neurons, 29 showed a slow, irregular firing pattern (< or = 2.5 spikes/s), 24 generated a fast continuous firing pattern (> or = 2.5 spikes/s), and 6 cells displayed a bursting pattern of activity consisting of alternating periods of spike discharge and quiescence. None of the bursting cells exhibited regenerative endogenous potentials (plateau potentials). On the contrary, in four of these cells, the bursting activity was clearly due to patterned synaptic activity. 5. The cultured OT cells responded to exogenous gamma-aminobutyric acid (GABA) and muscimol with a hyperpolarization and an increase in membrane conductance. These effects still were observed in the presence of TTX, indicating that they were due to direct activation of GABA receptors in the cells. The GABA-induced response was mediated by GABAA receptors because it was blocked by bicuculline, but not by GABAB receptors, because baclofen and hydroxysaclofen had no effect on membrane potential and input resistance. 6. OT neurons responded to exogenous glutamate, quisqualate, and kainate with a depolarization concomitant with an increase in membrane conductance. N-methyl-D-aspartate depolarized the cells in Mg(2+)-free medium. These effects were observed in the presence of TTX, suggesting that OT cells expressed ionotropic glutamate receptors. Trans-(1S,3R)-1-amino-1,3-cyclopentane-dicarboxylic acid and (+/-)-alpha-amino-4-carboxymethylphenylglycine had no effect on OT cells, thus excluding the presence of metabotropic glutamate receptors. 7. Taken together, our observations demonstrate that hypothalamic slice cultures from 4- to 6-day-old rat neonates contain well-differentiated OT neurons that display electrical properties similar to those shown by adult neurons in vitro. Such cultures provide a reliable model to investigate membrane properties of adult OT neurons and a useful means to study the long-term modulation of their electrical behaviour by various agents known to affect OT cells in vivo.

Action Potentials↗
PubMed1994

Evidence that TRH controls prolactin release from rat lactotrophs by stimulating a calcium influx.

Prolactin (PRL) release and intracellular free calcium concentration [Ca2+]i were measured in two populations of normal rat lactotrophs (light and heavy fractions) in culture. Spontaneous PRL release of heavy fraction cells was more sensitive to dihydropyridines (DHPs; Bay K 8644 and nifedipine) when compared to the light fraction lactotrophs. The stimulatory effect of thyrotropin-releasing hormone (TRH) on PRL release from heavy fraction cells was inhibited by Cd2+ and mimicked by Bay K 8644. Indo-1 experiments revealed that TRH-increased [Ca2+]i was reversibly inhibited by Cd2+. In a Ca(2+)-free EGTA-containing medium, TRH did not modify [Ca2+]i.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1993

Mechanism of the prolactin rebound after dopamine withdrawal in rat pituitary cells.

To study the mechanism underlying the effect of dopamine withdrawal on prolactin release, continuous perfusion experiments were performed on rat lactotroph-enriched primary cultures. Removal of dopamine (10(-7) M) after a short-term application (15 min) produced a rebound of prolactin secretion, which was enhanced by pretreatment of the cell culture with 17 beta-estradiol (10(-8) M for 48 h). Ca2+ channel blockade by Co2+ (1 mM) abolished the rebound in prolactin release. An increase in intracellular adenosine 3',5'-cyclic monophosphate by either forskolin (5 microM) or 3-isobutyl-1-methylxanthine (100 microM) enhanced the prolactin rebound after dopamine withdrawal. Application of thyrotropin-releasing hormone (10(-7) M) increased the prolactin rebound after dopamine withdrawal with a maximum effect obtained by commencing treatment immediately after removal of dopamine. Pretreatment of cell cultures with pertussis toxin (100 ng/ml, for 10 h) totally abolished the effects of dopamine on prolactin secretion. The dopamine agonist bromocriptine (10(-9) M) significantly decreased prolactin secretion, but no rebound effect was observed after its removal. We conclude that the rebound of prolactin release after dopamine treatment involves the influx of Ca2+.

Animals↗
PubMed1992

Dialysis of lactotropes with antisense oligonucleotides assigns guanine nucleotide binding protein subtypes to their channel effectors.

This article describes a new approach for determining the role of endogenous guanine nucleotide binding (G) protein subunits in signal transduction. Sequential patch-clamping was applied to BSA gradient-enriched cultured lactotropes from lactating rats, first to dialyze antisense oligodeoxyribonucleotides (AS) directed against G alpha protein mRNAs and 48 h later to record ion-current responses to the PRL release inhibitor, dopamine. The effectiveness and specificity of action of six types of AS were determined by their effects on the in vitro translation of alpha o, alpha i1, alpha i2, alpha i3, and alpha s. The specificity of AS could be enhanced by replacing guanine by cytosine bases within the center core of AS and by maximizing the number of mismatches against nontargeted mRNAs within the extremities of AS. A total of 59 out of 240 cells could be investigated using the sequential patch clamp procedure in the absence of antibiotics. The typical decrease of the voltage-activated calcium current in response to 10 nM dopamine was diminished or abolished by AS, in correlation with the inhibition of in vitro translation of the alpha o subunit. The typical increase of the voltage-activated potassium current in response to dopamine was abolished by AS directed against alpha i3 but not alpha o mRNA. Control experiments showed that culture conditions or loss of receptor affinity for dopamine were not responsible for the loss of response. The results suggest that dopamine D2 receptors are linked via alpha o to calcium channels and via alpha i3 to potassium channels.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Chronic stimulation of D2 dopamine receptors specifically inhibits calcium but not potassium currents in rat lactotrophs.

The present study examines the effect of chronic dopamine treatment, known to inhibit prolactin release from anterior pituitary, on two Ca2+ and K+ currents in cultured rat lactotrophs. K+ and Ca2+ currents were recorded using the whole-cell mode of the patch-clamp technique. The two types of voltage-dependent Ca2+ currents are called SD and FD (slowly deactivating and fast deactivating current component, respectively) and the two types of voltage-dependent K+ currents, IA and IK. All current types were isolated by tail current analysis. The amplitude of both normalized calcium components depended on the length of the culture (n = 48) while normalized amplitudes of both potassium currents remained constant (n = 9). Incubation of cells during 72 h with 50 microM of Actinomycin D, an inhibitor of mRNA synthesis, suggested that this increase in Ca2+ currents involved the synthesis of proteins. Long-lasting D2 receptor stimulation (8 days; 10 nM RU 24213) prevented this selective effect through activation of a pertussis toxin-sensitive G protein. We also examined whether cyclic adenosine-3',5'-cyclic-monophosphate (cyclic AMP) or Ca2+/phospholipid-dependent protein kinase (protein kinase C) could affect this development of channel activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Cervical rotation flaps for midface resurfacing.

The midface has long served as a focus for creativity in surgical reconstruction. Full-thickness skin grafts, split-thickness grafts, and distal flaps have long been used to attempt to reduplicate existing anatomy in this area. Recent reconstruction efforts have focused on the creative use of microvascular free flaps for this purpose. This article reports on the use of extensively developed regional rotation flaps as an excellent reconstructive modality for use in this area of the face. The details of surgical incisional planning are given. The nuances of surgical creation of these flaps and their rotation and suspension into place are given. The cases we have done using this technique for the past 3 years are reviewed. Our present indications for use of these flaps and their limitations are given.

Adult↗
PubMed1991

Physiological characterization of two functional states in subpopulations of prolactin cells from lactating rats.

1. Lactotroph cells from lactating female rat pituitary glands were dissociated, separated and enriched on a continuous gradient of bovine serum albumin at unit gravity. Two lactotroph subpopulations were observed in the light (F(3-5)) and the heavy (F(7-9)) fractions of the gradient. Both populations were maintained for at least 6 days in culture before experiments were performed. 2. Patch-clamp recordings, in the whole-cell mode, were performed on both lactotroph subpopulations in order to measure passive membrane properties and Ca2+ currents. Resting membrane potential as well as membrane capacitance values were found to be lower in light fraction cells. The two components of Ca2+ currents, called fast and slow deactivating (FD and SD) currents, were present with different proportions in each subpopulation; the ratio of current amplitudes, SD/FD, was 2.42 +/- 0.41 (n = 18) in light fraction cells and 1.17 +/- 0.27 (n = 17) in heavy fraction cells (P less than 0.02). 3. Reverse haemolytic plaque assay showed that in the light and heavy fractions, 68 and 47% of the lactotroph cells, respectively, were secreting. Population analysis of the plaque areas revealed a bimodal frequency distribution of plaque sizes consisting of small (1500 microns 2) and large plaques (3995 microns 2). A majority of light fraction cells produced large plaques whereas most of the heavy fraction cells produced small plaques. 4. Perifusion experiments performed on enriched prolactin cells showed that (1) basal prolactin (PRL) release was higher in light fraction than in heavy fraction cells, (2) the dopamine (10(-8)M)-induced inhibition of PRL release was greater in light fraction cells (86 +/- 15%) than in heavy fraction cells (41 +/- 21%), and (3) the thyrotrophin-releasing hormone (TRH, 10(-8)M)-induced increase of PRL release was 150 +/- 60% in light fraction versus 330 +/- 82% in heavy fraction cells. 5. Current-clamp recordings were performed using the intracellular technique. Lactotrophs were categorized according to their electrophysiological response following application of dopamine or TRH (both 10(-8)M). In the light fractions, the majority of the cells tested were hyperpolarized by dopamine (68%), whereas only 7% were depolarized by TRH application. In the heavy fractions, most of the cells (63%) responded to TRH application, while only 13% were dopamine sensitive. 6. Cytosolic free Ca2+ concentration ([Ca2+]i) measurements with the fluorescent probe Indo-1 revealed two lactotroph subtypes. Most cells in the light fractions (sixteen of twenty-two tested cells) exhibited an unstable level of [Ca2+]i with values fluctuating between 114.1 +/- 34.3 and 221 +/- 50 nM (mean +/- S.D.). Application of dopamine or of the D2 receptor agonist RU 24213 (10(-8)M) resulted in the disappearance of these fluctuations and in an accompanying decrease in basal [Ca2+]i level.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Different expression of the two dopaminergic D2 receptors, D2415 and D2444, in two types of lactotroph each characterised by their response to dopamine, and modification of expression by sex steroids.

Dopamine inhibits prolactin liberation acting via the D2 type receptor. Two different electrophysiological responses to dopamine have been shown to characterise two types of lactotroph isolated from the lactating female rat. It is now known that differential splicing of the pre-messenger RNA coding for the D2 receptor leads to the production of two D2 subtypes, D2(415) and D2(444). These subtypes differ in the region which is believed to be responsible for the binding of G proteins, and could thus lead to the activation of different intracellular second messenger systems. Here we show that the pre-messenger RNA for the D2 receptor is differentially spliced in such a way that the ratio D2(415)/D2(444) is significantly different (2.91 +/- 0.6 vs 1.29 +/- 0.14) between two populations of lactotrophs, each enriched in cells showing one type of response to DA. We further show that the ratio D2(415)/D2(444) can be changed by treatment of prolactin cells in primary culture with progesterone or testosterone. Estrogen did not change the ratio, but diminished the total amount of D2 cDNA. Regulation of differential splicing by sex steroids could provide a mechanism for modifying lactotroph responsiveness to DA in different physiological situations.

Animals↗
PubMed1990

Evidence for a relationship between lactotroph heterogeneity and physiological context.

Two lactotroph subpopulations differing in their functional properties have previously been isolated from lactating female rats. It was found that adult female rats at proestrus similarly yield two subpopulations of lactotrophs, whereas those from adult female rats at metestrus show different properties and in males only one functional population is found. As the period of lactation and proestrus are characterised by high circulating estrogen levels, estrogen is considered to modify lactotroph properties in the female, and as the functional differences concern the lactotroph responsiveness to TRH, it is supposed that lactotroph heterogeneity provides a means for particular secretory patterns during lactation and at proestrus.

Animals↗
PubMed1990

A guanine nucleotide-binding protein mediates the inhibition of voltage-dependent calcium currents by dopamine in rat lactotrophs.

The present study examines the effect of dopamine (DA), known to inhibit prolactin (PRL) release, on voltage-activated calcium currents in identified rat lactotrophs. Two types of voltage-dependent Ca2+ currents were recorded using the whole-cell mode of the patch-clamp technique. Both were reversibly inhibited by DA application. The inhibitory action of DA was reduced by (i) sulpiride (D2 antagonist), (ii) preincubation of the cells with pertussis toxin (PTX), and (iii) inclusion of guanosine 5'-O-(2-thiodiphosphate) (GDP-beta-S) in the pipette solution, whereas it was potentiated by guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S). This DA-induced response could not be overcome by changing the adenosine 3',5'-cyclic monophosphate level. These findings suggest that DA can inhibit Ca2+ entry through voltage-activated Ca2+ channels via a PTX-sensitive G protein(s) pathway thereby affecting PRL release from rat lactotrophs.

Animals↗
PubMed1990

Sodium and calcium currents in action potentials of rat somatotrophs: their possible functions in growth hormone secretion.

We report that both Na+ and Ca2+ currents are involved in the action potentials and in the hormone release from rat somatotrophs in primary culture. Single somatotrophs were identified by reverse hemolytic plaque assay (RHPA) and transmembrane voltage and currents were recorded using the whole-cell mode of the patch-clamp technique. Somatotrophs displayed a mean resting potential of -80mV and an average input resistance of 5.7G omega. Most of the cells showed spontaneous or evoked action potentials. Single action potentials or the initial spike in a burst were characterized by their high amplitude and short duration. Tetrodotoxin (TTX, 1 microM) blocked single action potentials and the initial spikes in a burst, whereas action potentials of long duration and low amplitude persisted. Cobalt (2 mM) plus TTX (1 microM) blocked all the action potentials. Voltage-clamp experiments confirmed the presence of both a TTX-sensitive Na+ current and Co2(+)-sensitive Ca2+ currents. TTX or Na(+)-free medium slightly decreased the basal release of GH but did not markedly modify hGRF-stimulated GH release. However, Co2+ (2 mM), which partially decreased the basal release, totally blocked hGRF-stimulated release. We conclude that (1) Na+ currents which initiate rapid action potentials may participate in spontaneous GH release; (2) Ca2+ currents, which give rise to long duration action potentials and membrane voltage fluctuation, are probably involved in both basal and hGRF-stimulated GH releases.

Action Potentials↗