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At least 73 records · Page 4Linked to original sources
PubMed2000

Effects of ATP on intracellular calcium dynamics of the perineurium of peripheral nerve bundles.

Adenosine-5'-triphosphate (ATP) released from damaged cells can affect functions of adjacent cells. Injuries of peripheral tissue stimulate nerves, but effect of ATP on the nerve bundles is still speculative. Peripheral nerves are surrounded by perineurium, therefore the response of perineurium may be a first event of nerve stimulation at tissue injuries. The aim of the present study is to clarify whether the perineurium responds to ATP. To this end, we analyzed the dynamics of the intracellular calcium concentration ([Ca2+]i) of perineurial cells by confocal microscopy. ATP induced a [Ca2+]i increase of perineurial cells. Ca2+ channel blockers and removing of extracellular Ca2+, but not thapsigargin pretreatment, abolished ATP-induced [Ca2+]i dynamics. This indicated that the [Ca2+]i increase was due to an influx of extracellular Ca2+. Adenosine-5'-diphosphate also elicited an increase of [Ca2+]i, but P1 receptor agonists had few effects on [Ca2+]i dynamics. Suramin (an antagonist of P2X and P2Y receptors) totally inhibited ATP-induced [Ca2+]i dynamics, but reactive blue 2 (a P2Y receptor antagonist) did not. Uridine-5'-triphosphate (a P2Y receptor agonist) induced no significant change in [Ca2+]i, but alpha,beta-methylene ATP (a P2X receptor agonist) caused a [Ca2+]i increase. In conclusion, perineurial cells respond to extracellular ATP mainly via P2X receptors.

Adenosine↗
PubMed2001

Effects of ATP on intracellular calcium dynamics of neurons and satellite cells in rat superior cervical ganglia.

Adenosine 5'-triphosphate (ATP) which is released from neuronal and non-neuronal tissues interacts with cell surface receptors to produce a broad range of physiological responses. The present study addressed the issue of whether the cells of the superior cervical ganglia (SCG) respond to ATP. To this end, the dynamics of the intracellular calcium ion concentration ([Ca2+]i) of neurons and satellite cells in intact SCG was analyzed by laser scanning confocal microscopy. ATP produced an increase of [Ca2+]i in both neurons and satellite cells; initially, ATP elicited [Ca2+]i increase in satellite cells and, subsequently, a [Ca2+]i change in neurons was observed. P1 purinoceptor agonists had no effect on this process, but P2 purinoceptor agonists induced [Ca2+]i increase and suramin totally inhibited ATP-induced [Ca2+]i dynamics in both neurons and satellite cells. In satellite cells, Ca2+ channel blockers and the removal of extracellular Ca2+, but not thapsigargin pretreatment, abolished ATP-induced [Ca2+]i dynamics. In contrast, thapsigargin pretreatment abolished ATP-induced [Ca2+]i dynamics in neurons. Reactive blue-2 inhibited the ATP-induced reaction on neurons alone. Uridine 5'-triphosphate caused a [Ca2+]i increase in neurons and alpha,beta-methylene ATP caused a [Ca2+]i increase in satellite cells. We concluded that neurons respond to extracellular ATP mainly via P2Y purinoceptors and that satellite cells respond via P2X purinoceptors.

Adenosine Triphosphate↗
PubMed2006

A buffering SERCA pump in models of calcium dynamics.

Many cells use oscillations in calcium concentration to transmit messages. The oscillations largely result from an influx of calcium into the cytosol from the endoplasmic reticulum (ER), followed by an efflux of calcium from the cytosol back into the ER. The sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) pump pumps calcium into the ER. It binds calcium on the cytosolic side and releases it on the ER side and in the delay between binding and release, calcium is buffered by the pump. We developed a model of a buffering SERCA pump and investigated whether including this in a model of calcium oscillations has any significant effects. We found that the oscillations produced when using the SERCA pump, which does not buffer calcium, have a larger amplitude and a slightly smaller period than when using the buffering SERCA pump. We show that the buffering SERCA pump shows adaptation to a stimulus, and we demonstrate that, by using a bidirectional SERCA pump, we are able to eliminate futile cycling of calcium between the cytosol and ER when the cell is at rest.

Biological Clocks↗
PubMed2000

Monitoring presynaptic calcium dynamics in projection fibers by in vivo loading of a novel calcium indicator.

Fluorometric calcium measurements have revealed presynaptic residual calcium (Ca(res)) to be an important regulator of synaptic strength. However, in the mammalian brain, it has not been possible to monitor Ca(res) in fibers that project from one brain region to another. Here, we label neuronal projections by injecting dextran-conjugated calcium indicators into brain nuclei in vivo. Currently available dextran conjugates distort Ca(res) due to their high affinity for calcium. Therefore, we synthesized a low-affinity indicator, fluo-4 dextran, that can more accurately measure the amplitude and time course of Ca(res). We then demonstrate the utility of fluo-4 dextran by measuring Ca(res) at climbing fiber presynaptic terminals. This method promises to facilitate the study of many synapses in the mammalian CNS, both in brain slices and in vivo.

Aniline Compounds↗
PubMed1996

Calcium dynamics and homeostasis in a mathematical model of the principal cell of the cortical collecting tubule.

Calcium (Ca) dynamics are incorporated into a mathematical model of the principal cell in the cortical collecting tubule developed earlier in Strieter et al. (1992a. Am. J Physiol. 263:F1063-1075). The Ca components are modeled after the Othmer-Tang model for IP(3)-sensitive calcium channels (1993, in Experimental and Theoretical Advances in Biological Pattern Formation, 295-319). There are IP(3)-sensitive Ca channels and ATP-driven pumps on the membrane of the endoplasmic reticulum. Calcium enters the cell passively down its electrochemical gradient. A Ca pump and Na/Ca exchange in the basolateral membrane are responsible for the extrusion of cytoplasmic calcium. Na/Ca exchange can also operate in reverse mode to transport Ca into the cell. Regulatory effects of cytoplasmic Ca on the apical Na channels are modeled after experimental data that indicate apical Na permeability varies inversely with cytoplasmic Ca concentration. Numerical results on changes in intracellular Ca caused by decreasing NaCl in the bath and the lumen are similar to those from experiments in Bourdeau and Lau (1990. Am. J Physiol. 258:F1497-1503). This match of simulation and experiment requires the synergistic action of the Na/Ca exchanger and the Ca regulated apical Na permeability. In a homogeneous medium, cytoplasmic Ca becomes oscillatory when extracellular Na is severely decreased, as observed in experiments of cultured principal cells (Koster, H., C. van Os and R. Bindels. 1993. Kidney Int.43:828-836). This essentially pathological situation arises because the hyperpolarization of membrane potential caused by Na-free medium increases Ca influx into the cell, while the Na/Ca exchanger is inactivated by the low extracellular Na and can no longer move Ca out of the cell effectively. The raising of the total amount of intracellular Ca induces oscillatory Ca movement between the cytoplasm and the endoplasmic reticulum. Ca homeostasis is investigated under the condition of severe extracellular Ca variations. As extracellular Ca is decreased, Ca regulation is greatly impaired if Ca does not regulate apical ionic transport. The simulations indicate that the Na/Ca exchanger alone has only limited regulatory capacity. The Ca regulated apical sodium or potassium permeability are essential for regulation of cytoplasmic Ca in the principal cell of the cortical collecting tubule.

Calcium↗
PubMed1999

Cameleon calcium indicator reports cytoplasmic calcium dynamics in Arabidopsis guard cells.

Cytoplasmic free calcium ([Ca2+]cyt) acts as a stimulus-induced second messenger in plant cells and multiple signal transduction pathways regulate [Ca2+]cyt in stomatal guard cells. Measuring [Ca2+]cyt in guard cells has previously required loading of calcium-sensitive dyes using invasive and technically difficult micro-injection techniques. To circumvent these problems, we have constitutively expressed the pH-independent, green fluorescent protein-based calcium indicator yellow cameleon 2.1 in Arabidopsis thaliana (Miyawaki et al. 1999; Proc. Natl. Acad. Sci. USA 96, 2135-2140). This yellow cameleon calcium indicator was expressed in guard cells and accumulated predominantly in the cytoplasm. Fluorescence ratio imaging of yellow cameleon 2.1 allowed time-dependent measurements of [Ca2+]cyt in Arabidopsis guard cells. Application of extracellular calcium or the hormone abscisic acid (ABA) induced repetitive [Ca2+]cyt transients in guard cells. [Ca2+]cyt changes could be semi-quantitatively determined following correction of the calibration procedure for chloroplast autofluorescence. Extracellular calcium induced repetitive [Ca2+]cyt transients with peak values of up to approximately 1.5 microM, whereas ABA-induced [Ca2+]cyt transients had peak values up to approximately 0.6 microM. These values are similar to stimulus-induced [Ca2+]cyt changes previously reported in plant cells using ratiometric dyes or aequorin. In some guard cells perfused with low extracellular KCl concentrations, spontaneous calcium transients were observed. As yellow cameleon 2.1 was expressed in all guard cells, [Ca2+]cyt was measured independently in the two guard cells of single stomates for the first time. ABA-induced, calcium-induced or spontaneous [Ca2+]cyt increases were not necessarily synchronized in the two guard cells. Overall, these data demonstrate that that GFP-based cameleon calcium indicators are suitable to measure [Ca2+]cyt changes in guard cells and enable the pattern of [Ca2+]cyt dynamics to be measured with a high level of reproducibility in Arabidopsis cells. This technical advance in combination with cell biological and molecular genetic approaches will become an invaluable tool in the dissection of plant cell signal transduction pathways.

Amino Acid Sequence↗
PubMed1994

A model of calcium dynamics in cardiac myocytes based on the kinetics of ryanodine-sensitive calcium channels.

The ryanodine-sensitive calcium channels are pivotal to signal transduction and cell function in many cell types, including cardiac myocytes. In this paper a kinetic model is proposed for these channels. In the model there are two Ca regulatory sites on the channel protein, one positive and the other negative. Cytoplasmic Ca binds to these regulatory sites independently It is assumed that the binding of Ca to the positive site is a much faster process than binding to the negative site. At steady state, the channel opening as a function of the Ca concentration is a bell-shaped curve. The model predicts the adaptation of channels to constant Ca stimulus. When this model is applied to cardiac myocytes, it predicts excitability with respect to Ca perturbations, smoothly graded responses, and Ca oscillations in certain pathological circumstances. In a spatially distributed system, traveling Ca waves in individual myocytes exist under certain conditions. This model can also be applied to other systems where the ryanodine-sensitive channels have been identified.

Adaptation, Physiological↗
PubMed1998

Intracellular calcium dynamics in mouse model of myocardial stunning.

Intracellular calcium (Cai2+) and left ventricular (LV) function were determined in the coronary-perfused mouse heart to study Cai2+-related mechanisms of injury from myocardial ischemia and reperfusion. Specifics for loading of the photoprotein aequorin into isovolumically contracting mouse hearts under constant-flow conditions are provided. The method allows detection of changes in Cai2+ on a beat-to-beat basis in a model of myocardial stunning and permits correlation of interventions that regulate Ca2+ exchange with functional alterations. Twenty-three coronary-perfused mouse hearts were subjected to 15 min of ischemia followed by 20 min of reperfusion. In 13 hearts, the perfusate included the calmodulin antagonist W7 (10 microM) to inhibit Ca(2+)-calmodulin-regulated mechanisms. Peak Cai2+ was 0.77 +/- 0.03 microM in the control group and was unaffected by W7 at baseline. Ischemia was characterized by a rapid decline in LV function, followed by ischemic contracture, accompanied by a gradual rise in Cai2+. Reperfusion was characterized by an initial burst of Cai2+ and a gradual recovery to nearly normal systolic Cai2+ while LV pressure recovered to 55% after 20 min of reperfusion (stunned myocardium). These results in the mouse heart confirm that stunning does not result from deficiency of Cai2+ but rather from a decreased myofilament responsiveness to Cai2+ due to changes in the myofilaments themselves. In hearts perfused with W7, the rise in Cai2+ during ischemia was significantly attenuated, as was the magnitude of mean Cai2+ during early reflow. Ischemic contracture was abolished or delayed. Hearts perfused with W7 showed significantly improved recovery of LV pressure, rate of contraction, and rate of relaxation. Diastolic Cai2+ was increased in control hearts during stunning but returned to baseline in hearts perfused with W7. Simultaneous assessment of Cai2+ and LV function demonstrates that calmodulin-regulated mechanisms may contribute to the pathogenesis of myocardial stunning in the mouse heart.

Animals↗
PubMed1988

Spatially resolved calcium dynamics of mammalian Purkinje cells in cerebellar slice.

Microfluorometric imaging was used to study the correlation of intracellular calcium concentration with voltage-dependent electrical activity in guinea pig cerebellar Purkinje cells. The spatiotemporal dynamics of intracellular calcium concentration are demonstrated during spontaneous and evoked activity. The results are in agreement with hypotheses of dendritic segregation of calcium conductances suggested by electrophysiological experiments. These in vitro slice fluorescence imaging methods are applicable to a wide range of problems in central nervous system biochemical and electrophysiological functions.

Animals↗
PubMed1996

The effect of age on calcium dynamics in rat brain in vivo.

Different biochemical changes accompany development and ageing. However, the time frame for a particular biochemical change may be different than that for another. In the ageing brain, neurological deficits related to ageing have been suggested to be due to a breakdown of calcium (Ca++) homeostasis, and an increase in intracellular Ca+2. To understand the dynamics of calcium in CNS in relation to normal ageing, we have determined 45Ca++ uptake by the different regions of brain in young (3 months) and aged (15 months) rats after a single i.v. injection of 90 microCi/kg 45Ca++. In the a-ed group, the initial plasma contents of the tracer were higher than in the young rats (3 months). The mean values of uptake (as fraction of injected dose/g x 10(5)) of 45Ca at different sites were higher in the 15-month-old animals than in the young rats (viz. cortex: 50 vs. 26; hippocampus: 90 vs. 46 and putamen: 54 vs. 30 at 120 min). Computer simulation of the uptake data indicated that the values of most of the fractional transport rates were similar or lower in the aged compared to the controls. However, the steady state ratio of intra/extracellular Ca++ was higher in the 15-month-old animals, suggestive of intracellular accumulation of Ca++.

Aging↗
PubMed1983

Calcium dynamics at a plastic synapse in Aplysia.

Because certain primitive behavioral responses in the large sea snail Aplysia have recently been linked to neurophysiological events at a synaptic level, special interest attaches to the role played by calcium ions at such synapses. Using an extended version of the model applied earlier to trace the flow of energy and information through a ganglion of the medicinal leech (Triffet & Green, 1980), the authors investigate the electropotential effects of small transient localized changes in the calcium concentration near the inner membrane surface of a neuron in the resting state. When this state is well below the firing threshold, changes in Ca2+ concentration less than 10(-8)M are shown to result only in low-level harmonic background oscillations. When the potential of the neurons is closer to threshold, however, and/or the Ca2+ concentration is of the order of 10(-8)M, easily recognizable graded potentials appear, and these grow into firing peaks when the calcium concentration is increased still further. Though no attempt is made to deal with the amplification effects dependent on calcium-vesicle interactions and the related release of transmitter molecules, a unified mechanism for the underlying calcium ion dynamics is proposed. Graded potentials of increasing size are associated with a progressive localized thickening of the inner and outer Debye layers. Moreover, the transverse and longitudinal calcium currents set up in such regions prove adequate to account for both the depletion of Ca2+ ions necessary to achieve habituation, and the increase in their concentration required for sensitization.

Action Potentials↗
PubMed1992

Calcium dynamics in neurons treated with toxic and non-toxic concentrations of glutamate.

Intracellular calcium concentration ([Ca2+]i) dynamics were simultaneously monitored in multiple cultured rat neurons loaded with Fluo-3 and continuously stimulated with glutamate (GLU). Three response types were observed: 10 microM GLU caused an initial transient increase in [Ca2+]i; 20 microM a biphasic response characterized by a 150-350 s 'calcium trough' between peaks; and 40 microM an initial sustained increase in [Ca2+]i. Neurons in calcium-free medium treated with 40 microM GLU showed only an initial transient increase in [Ca2+]i, demonstrating the dependence of sustained secondary increases in [Ca2+]i on extracellular calcium sources. We observed synchronized responses of multiple neurons within a given culture well, after GLU treatment, supporting the hypothesis that sustained influx of extracellular calcium may be stimulated by depletion of intracellular calcium and/or the release of endogenous excitatory amino acids.

Aniline Compounds↗
PubMed1996

Spatial organization of calcium dynamics in growth cones of sensory neurones.

The concentration of calcium ions in the cytosol ([Ca2+]i) has a dominant influence on neuronal development. A [Ca2+]i rise can, depending on the amplitude and location, promote outgrowth or dramatically inhibit it. We have used the fluorescent calcium indicators Fura-2 and Fura-2 dextran to measure [Ca2+]i dynamics in sensory neurones from the adult rat. [Ca2+]i was low and uniform in advancing growth cones, even during specific behaviours such as protrusion, filling and consolidation. A brief train of action potentials caused [Ca2+]i to rise at the extreme leading edge of the growth cone. [Ca2+]i changes in more proximal regions of the growth cone were much smaller. This spatially organized [Ca2+]i change, which may result from a concentration of calcium channels at the growth cone leading edge, is likely to function in spontaneously active regenerating axons in vivo to specifically activate calcium-dependent processes at the growth cone tip.

Animals↗
PubMed2000

The relationship between pulsatile secretion and calcium dynamics in single, living gonadotropin-releasing hormone neurons.

It is well established that pulsatile release of GnRH regulates the reproductive axis, but little is known about the mechanisms underlying this pulsatility. Recent findings that GT1 cells, a line derived from the mouse embryonic hypothalamus, release GnRH in a pulsatile manner indicates that this rhythmic activity is an intrinsic property of GnRH neurons. In several attempts to uncover the intracellular basis for this pulsatile phenomenon, it was revealed that intracellular calcium concentrations change in a rhythmic fashion in GnRH neurons and that cellular depolarization, which triggers a secretory event, is associated with profound calcium changes in the cells. These findings raised the intriguing possibility that periodic alterations in intracellular calcium concentrations may underlie the phenomenon of pulsatile secretion in GnRH neurons. To address this, we first adapted the use of FM1-43 fluorescence to monitor changes of secretion in individual GT1-7 cells and then combined this approach with simultaneous measurement ofintracellular free calcium ([Ca2+]i, fura 2 method). In initial validation experiments, we found that stimulation of exocytosis with K+ (75 mM) or N-methyl-D-aspartate (NMDA, 100 microM) predictably evoked dynamic increases of both FM1-43 and fura 2 fluorescence. Later measurement of calcium dynamics and exocytotic activity in unstimulated cells revealed that [Ca2+]i underwent transitions from quiescence to high oscillatory behavior, and that these shifts were frequently associated with exocytotic events. Moreover, these calcium oscillatory transitions and associated changes in secretory activity occurred synchronously among most adjacent cells and at a frequency similar to that reported for pulsatile release of GnRH by entire cultures of GnRH neurons. Taken together, these results indicate that the intrinsic secretory pulsatility of GnRH neurons appears to be a consequence of coordinated, periodic changes in the pattern of calcium oscillations within individual cells.

Animals↗
PubMed1999

Effects of AIF4- and ATP on intracellular calcium dynamics of crypt epithelial cells in mouse small intestine.

Previous digital imaging analysis of intracellular calcium ion ([Ca2+]i) dynamics in the crypt of the small intestine showed little response by most columnar cells to cholinergic and adrenergic agonists. The objective of the present study was to demonstrate whether G-protein activators and other transmitters elicit [Ca2+]i changes in crypt cells. We used digital imaging to analyze spatiotemporal dynamics of [Ca2+]i in Fura-2/AM-loaded isolated crypts of mouse duodenum and ileum. A1F4- increased [Ca2+]i in crypt columnar cells. In many cases, we observed [Ca2+]i oscillations, which were synchronized throughout the entire crypt. The oscillations were blocked by octanol. ATP, but not adenosine, caused a [Ca2+]i increase in middle crypt-regions of the duodenum and upper regions of the ileum, and the [Ca2+]i wave propagated towards the crypt bottom. The ATP-induced [Ca2+]i increase was prevented by pretreatment with thapsigargin or suramin, but not by La3+ or an extracellular Ca(2+)-free environment. Neither dopamine, 5-hydroxytryptamine (5-HT), histamine, vasoactive intestinal peptide, substance P. cholera toxin, nor guanylin had significant effects. The [Ca2+]i dynamics of Paneth cells were independent of the AlF4(-)-induced synchronous oscillations of columnar cells and of the ATP-induced [Ca2+]i wave. In conclusion, crypt columnar cells have [Ca2+]i-dependent intracellular signaling mechanisms that are linked with G proteins, and by which the cells communicate with each other. ATP elicited [Ca2+]i mobilization from columnar cells via P2 receptors, although some regional differences were noted between the duodenum and ileum.

Adenosine Triphosphate↗
PubMed1994

Calcium dynamics during starfish oocyte maturation and fertilization.

Intracellular free calcium levels in starfish oocytes have been monitored during meiotic maturation and fertilization using calcium-sensitive fluorescent dyes combined with confocal laser scanning microscopy or fura ratioing techniques. In time-lapse analyses of prophase-arrested and maturing oocytes, calcium transients were elicited by inositol 1,4,5-trisphosphate (IP3), ryanodine, or caffeine, indicating that both the IP3-sensitive and IP3-insensitive receptors of the oocyte's calcium release channels could be stimulated to mobilize calcium ions. Fertilization also triggered a global calcium wave that appeared to travel faster around the cortex than through the center of the oocyte, and maturing oocytes developed normally after their fertilization-induced calcium waves had been imaged. Prophase-arrested specimens, on the other hand, did not undergo germinal vesicle breakdown or cleavage after displaying a fertilization-induced calcium transient throughout their cytoplasm and nucleus, confirming previous observations that calcium spikes are not sufficient to induce development in immature oocytes. In addition, although the calcium spikes triggered by sperm or caffeine reached similar normalized peak heights, fertilization-induced calcium waves in maturing oocytes tended to be more prolonged than the fertilization waves observed in prophase-arrested oocytes or the caffeine-triggered spikes elicited at any stage of maturation. Collectively, such findings suggest that the total amount of releasable calcium does not vary appreciably during maturation, but the patterns of the calcium transients can differ depending on the stage of maturation and/or the type of calcium-releasing agent. Possible artifacts affecting these findings are assessed, and the results are discussed relative to the functioning of calcium release pathways during starfish oocyte maturation and fertilization.

Animals↗
PubMed2005

Fluoxetine inhibits calcium-activated currents of salamander rod photoreceptor somata and presynaptic terminals via modulation of intracellular calcium dynamics.

PURPOSE: In order to isolate voltage-gated calcium currents in rods retaining intact axons and presynaptic terminals, it is first necessary to identify specific blockers of the large calcium-dependent chloride current, ICl(Ca), which obscures them. Based upon previous reports of its efficacy as an inhibitor of a volume regulated chloride channel (VRAC), a calcium-dependent chloride channel, and the cystic fibrosis transmembrane conductance regulator (CFTR), we investigated whether the serotonin reuptake inhibitor, fluoxetine hydrochloride, could act as a specific blocker for ICl(Ca) in salamander rod photoreceptor terminals, without affecting other aspects of rod physiology. METHODS: Intact rod photoreceptors retaining axons and presynaptic terminals were enzymatically dissociated from salamander retinae. Under whole cell voltage clamp, depolarization-induced whole cell currents were recorded in the presence and absence of fluoxetine (10 and 50 microM pipette concentration) administered via a puffing pipette or in the bath (25 microM). Changes in intracellular free calcium levels were monitored as changes in fura-2 fluorescence following brief depolarization with high K+ (50 and 100 mM) administered via a puffing pipette in the presence and absence of fluoxetine (4 and 10 microM) in the bath. RESULTS: When puffed onto cells, fluoxetine inhibited ICl(Ca) in a dose-dependent fashion (50 microM=96% reduction; 10 microM=14% reduction). In addition to the reduction in amplitude of ICl(Ca), 4 microM fluoxetine (pipette concentration) significantly reduced the duration of ICl(Ca) (48% reduction). Fluoxetine also suppressed the calcium-activated potassium current, IK(Ca), to similar extents (50 microM=75% reduction; 10 microM=23% reduction) when puffed onto cells. Preincubation of rods with 25 microM fluoxetine in the bath significantly reduced outward currents at both 0 mV, where ICl(Ca) is negligible because ECl is about 0 mV and the bulk of the current is carried by IK(Ca), and at +40 mV, where the current is a combination of ICl(Ca) and IK(Ca). Parallel calcium imaging experiments with fura-2 revealed that preincubation of rods in 10 microM fluoxetine virtually eliminated the normal rise in intracellular free calcium in somatic (99.6% reduction) and terminal (98% reduction) compartments following brief depolarization with high K+ (100 mM pipette concentration). Cells preincubated in 4 microM fluoxetine, a therapeutically relevant concentration, showed smaller but significant reductions in Ca2+ elevations in both somatic (66% reduction) and terminal (36% reduction) compartments and even more significant reductions in the duration of sustained calcium levels of the terminal compartment (50% reduction) following brief depolarization with high K+ (50 mM pipette concentration). CONCLUSIONS: We conclude that in addition to blocking ICl(Ca), fluoxetine inhibits IK(Ca). We further conclude that the inhibition of both of these currents is the consequence of inhibition of the normal sustained elevation in intracellular calcium following depolarization and initial calcium influx. Combined, the data suggest that fluoxetine may have multiple sites of action in rod photoreceptors instead of acting as a specific inhibitor of ICl(Ca).

Ambystoma↗
PubMed2025

Nongenomic Stimulatory Effect of T3 on Calcium Dynamics in GnRH Neurons via Integrin αVβ3.

Many clinical studies have identified correlations between thyroid dysfunction and reproductive issues, yet the underlying mechanisms behind this interaction remain poorly understood. In this study, we investigated the effect of triiodothyronine (T3) on the activity of gonadotropin-releasing hormone (GnRH) neurons, a key regulator of the central reproductive axis. Dual labeling confirmed that GnRH neurons express thyroid receptor (TR)α and integrin αVβ3 receptors mediating genomic and nongenomic effects of thyroid hormones, respectively. Using calcium imaging in an ex vivo model, we show that T3 induces a rapid and sustained increase of calcium oscillation frequency in GnRH neurons. No change in response was detected after application of T4. The T3 stimulatory effect was not inhibited by a TR-specific antagonist (1-850) but was mimicked by membrane-impermeable T3-BSA, indicating a mechanism independent of nuclear TR signaling. In contrast, the blockade of membrane αVβ3 integrins (with cilengitide) prevented the T3-induced increase in GnRH neurons calcium peak oscillation frequency. Further investigation using modulators of intracellular calcium and calcium entry revealed that binding to αVβ3 integrin can induce distinct calcium responses depending on the ligand, with T3 triggering a complex response involving multiple channels and calcium sources, possibly with compensatory mechanisms. In sum, these results demonstrate for the first time a direct effect of thyroid hormones on GnRH neuronal activity, with T3 stimulating calcium oscillations through the nongenomic αVβ3 integrin pathway. Understanding this thyroid-reproductive axis interaction will help clarify the mechanisms linking thyroid dysfunction to reproductive disorders and pave the way for targeted therapeutic interventions.

Animals↗