Calcium channels and pumps in cancer: changes and consequences

Increases in intracellular free Ca2+ play a major role in many cellular processes. The deregulation of Ca2+ signaling is a feature of a variety of diseases, and modulators of Ca2+ signaling are used to treat conditions as diverse as hypertension to pain. The Ca2+ signal also plays a role in processes important in cancer, such as proliferation and migration. Many studies in cancer have identified alterations in the expression of proteins involved in the movement of Ca2+ across the plasma membrane and subcellular organelles. In some cases, these Ca2+ channels or pumps are potential therapeutic targets for specific cancer subtypes or correlate with prognosis

Function of Cav3.2-T-type in human prostatic cancer cells

Le cancer de la prostate évolue progressivement vers l'androgéno-indépendance, stade incurable pour lequel la recherche biomédicale tente de déterminer de nouvelles cibles thérapeutiques. Cette évolution est marquée par une différenciation neuroendocrine des cellules prostatiques qui s'accompagne d'une surexpression de canaux calciques voltage-dépendants de type T (Cav3.2) responsable d'une altération de l'homéostasie calcique intracellulaire. L'objectif de cette thèse était donc de déterminer l'implication des Cav3.2 dans les cellules cancéreuses prostatiques humaines, et notamment dans les cellules neuroendocrines. Nos résultats montrent que, dans ces cellules non-excitables, les Cav3.2 participent à l'homéostasie calcique en permettant une entrée basale de calcium au potentiel de repos sans contribuer à une entrée capacitive de calcium provoquée par la vidange des réserves intracellulaires de calcium. De plus, ces canaux interviennent dans la sécrétion de la Phosphatase Acide Prostatique, un marqueur des cellules épithéliales prostatiques, dans les cellules neuroendocrines. Par ailleurs, nous montrons l'existence d'un couplage fonctionnel entre les Cav3.2 et les canaux de type BK qui constituent les principales conductances potassiques voltage-dépendantes des cellules prostatiques. Enfin, nos travaux mettent en évidence que ce couplage entre ces deux canaux participe à la prolifération cellulaire des cellules cancéreuses prostatiques. En conclusion, ce travail de thèse contribue à élargir les connaissances sur les canaux calciques de type T, Cav3.2 en particulier, et leur rôle au sein des cellules prostatiques cancéreuses humainesProstate cancer inevitably evolves towards an incurable androgen-independent stage for which biomedical research attempts to identity new therapeutic targets. This progression is characterized by a neuroendocrine differentiation of prostate cells accompanied by an overexpression of voltage-dependent T-type calcium channels (Cav3.2), responsible for an alteration of intracellular calcium homeostasis. The aim of this PhD thesis was to determine the involvement of Cav3.2 in human prostate cancer cells, mainly in neuroendocrine cells. Our results show that, in these non-excitable cells, Cav3.2 channels participate to basal calcium homeostasis by promoting calcium entry at resting membrane potential without contributing to the capacitative calcium entry triggered by calcium depletion from endoplasmic reticulum stores. ln addition, these channels are involved in the secretion of Prostatic Acid Phosphatase, a marker of prostatic epithelial cells, in neuroendocrine cells. Furthermore, we show that Cav3.2 channels are functionaly coupled to BK channels which constitute the main voltage-dependent potassium conductances in prostate cells. Finally, our work demonstrates that this coupling between both channels regulates prostate cancer cell proliferation. As a conclusion, this work contributes to the understanding of the role of Cav3.2 T-type calcium channels in human prostate cancer cells

Function of Cav3.2-T-type in human prostatic cancer cells

Le cancer de la prostate évolue progressivement vers l'androgéno-indépendance, stade incurable pour lequel la recherche biomédicale tente de déterminer de nouvelles cibles thérapeutiques. Cette évolution est marquée par une différenciation neuroendocrine des cellules prostatiques qui s'accompagne d'une surexpression de canaux calciques voltage-dépendants de type T (Cav3.2) responsable d'une altération de l'homéostasie calcique intracellulaire. L'objectif de cette thèse était donc de déterminer l'implication des Cav3.2 dans les cellules cancéreuses prostatiques humaines, et notamment dans les cellules neuroendocrines. Nos résultats montrent que, dans ces cellules non-excitables, les Cav3.2 participent à l'homéostasie calcique en permettant une entrée basale de calcium au potentiel de repos sans contribuer à une entrée capacitive de calcium provoquée par la vidange des réserves intracellulaires de calcium. De plus, ces canaux interviennent dans la sécrétion de la Phosphatase Acide Prostatique, un marqueur des cellules épithéliales prostatiques, dans les cellules neuroendocrines. Par ailleurs, nous montrons l'existence d'un couplage fonctionnel entre les Cav3.2 et les canaux de type BK qui constituent les principales conductances potassiques voltage-dépendantes des cellules prostatiques. Enfin, nos travaux mettent en évidence que ce couplage entre ces deux canaux participe à la prolifération cellulaire des cellules cancéreuses prostatiques. En conclusion, ce travail de thèse contribue à élargir les connaissances sur les canaux calciques de type T, Cav3.2 en particulier, et leur rôle au sein des cellules prostatiques cancéreuses humainesProstate cancer inevitably evolves towards an incurable androgen-independent stage for which biomedical research attempts to identity new therapeutic targets. This progression is characterized by a neuroendocrine differentiation of prostate cells accompanied by an overexpression of voltage-dependent T-type calcium channels (Cav3.2), responsible for an alteration of intracellular calcium homeostasis. The aim of this PhD thesis was to determine the involvement of Cav3.2 in human prostate cancer cells, mainly in neuroendocrine cells. Our results show that, in these non-excitable cells, Cav3.2 channels participate to basal calcium homeostasis by promoting calcium entry at resting membrane potential without contributing to the capacitative calcium entry triggered by calcium depletion from endoplasmic reticulum stores. ln addition, these channels are involved in the secretion of Prostatic Acid Phosphatase, a marker of prostatic epithelial cells, in neuroendocrine cells. Furthermore, we show that Cav3.2 channels are functionaly coupled to BK channels which constitute the main voltage-dependent potassium conductances in prostate cells. Finally, our work demonstrates that this coupling between both channels regulates prostate cancer cell proliferation. As a conclusion, this work contributes to the understanding of the role of Cav3.2 T-type calcium channels in human prostate cancer cells

Psychothérapie alcoologique de groupe selon le modèle cognitivo-comportemental

National audienc

A role for voltage gated T-type calcium channels in mediating “capacitative” calcium entry?

Calcium entry through plasma membrane calcium channels is one of the most important cell signaling mechanism involved in such diverse functions as secretion, contraction and cell growth by regulating gene expression, proliferation and apoptosis. The identity of plasma membrane calcium channels, the main regulators of calcium entry, involved in cell proliferation has been thus extensively sought. Among these, a calcium entry pathway called capacitative calcium entry (CCE), activated by calcium store depletion, is particularly important in non-excitable cells. Though this capacitative calcium entry is generally supposed to occur through TRP channels there is some evidence that voltage-dependent T-type calcium channels may contribute to calcium entry after store depletion. Here we show that though mibefradil, a T-type calcium channel blocker, is able to reduce capacitative calcium entry induced by either thapsigargin or ATP, this was not mimicked by any other T-type calcium channel inhibitors even in cells overexpressing ␣ 1H T-type calcium channels, leading us to conclude that T-type calcium channels are not responsible for the capacitative calcium entry observed in different cancer cell lines. On the contrary, we show that the action of mibefradil on capacitative calcium entry is due to an action on store-operated calcium channels

Variability of quantal NMDA to AMPA current ratio in nucleus tractus solitarii neurons

The ratio between AMPA and NMDA receptors is a key factor governing integrative and plastic properties of excitatory glutamatergic synapses. To determine whether the respective proportions of AMPA and NMDA receptors are similar or vary across a neuron's synapse, we analyzed the variability of NMDA and AMPA currents in quantal responses recorded from neurons located in the nucleus tractus solitarii. We found that the average NMDA to AMPA current ratio strongly differed between recorded neurons and that most of the intra-neuronal current ratio variability was attributable to fluctuations in NMDA current. We next performed computer simulations with a Monte Carlo model of a glutamatergic synapse to estimate the part of AMPA and NMDA currents fluctuations induced by stochastic factors. We found that NMDA current variability mainly resulted from strong channel noise with few influence of release variations. On the contrary, partly because of the presence of subconductance states, AMPA receptor channel noise was low and AMPA current fluctuations tightly reflected changes in the amount of glutamate released. We next showed that these two factors, channel noise and fluctuations in glutamate release, were sufficient to explain the observed variability of the NMDA to AMPA current ratio in quantal events recorded from the same neuron. We therefore concluded that the proportion of AMPA and NMDA receptors was similar, or roughly similar, across synapses onto the same target cell

Prochlorperazine Increases KCC2 Function and Reduces Spasticity after Spinal Cord Injury.

International audienceIn mature neurons, low intracellular chloride level required for inhibition is maintained by the potassium-chloride co-transporter KCC2. Impairment of Cl- extrusion following KCC2 dysfunction has been involved in many CNS disorders such as seizures, neuropathic pain or spasticity after a spinal cord injury (SCI). This makes KCC2 an appealing drug target for restoring Cl-homeostasis and inhibition in pathological conditions. In the present study, we screen the Prestwick Chemical Library® and identify conventional antipsychotics phenothiazine derivatives as enhancers of KCC2 activity. Among them, prochlorperazine hyperpolarizes the Cl- equilibrium potential in motoneurons of neonatal rats and restores the reciprocal inhibition after SCI. The compound alleviates spasticity in chronic adult SCI rats with an efficacy equivalent to the anti-spastic agent baclofen, and rescues the SCI-induced downregulation of KCC2 in motoneurons below the lesion. These preclinical data support prochlorperazine for a new therapeutic indication in the treatment of spasticity after SCI and neurological disorders involving a KCC2 dysfunction

Functional coupling between large-conductance potassium channels and Cav3.2 voltage-dependent calcium channels participates in prostate cancer cell growth

Summary It is strongly suspected that potassium (K+) channels are involved in various aspects of prostate cancer development, such as cell growth. However, the molecular nature of those K+ channels implicated in prostate cancer cell proliferation and the mechanisms through which they control proliferation are still unknown. This study uses pharmacological, biophysical and molecular approaches to show that the main voltage-dependent K+ current in prostate cancer LNCaP cells is carried by large-conductance BK channels. Indeed, most of the voltage-dependent current was inhibited by inhibitors of BK channels (paxillin and iberiotoxin) and by siRNA targeting BK channels. In addition, we reveal that BK channels constitute the main K+ channel family involved in setting the resting membrane potential in LNCaP cells at around −40 mV. This consequently promotes a constitutive calcium entry through T-type Cav3.2 calcium channels. We demonstrate, using single-channel recording, confocal imaging and co-immunoprecipitation approaches, that both channels form macromolecular complexes. Finally, using flow cytometry cell cycle measurements, cell survival assays and Ki67 immunofluorescent staining, we show that both BK and Cav3.2 channels participate in the proliferation of prostate cancer cells

CaV3.2 T-type Calcium Channels Are Involved in Calcium-dependent Secretion of Neuroendocrine Prostate Cancer Cells

International audienceBecause prostate cancer is, in its early stages, an androgen-dependent pathology, treatments aiming at decreasing testosterone plasma concentration have been developed for many years now. However, a significant proportion of patients suffer a relapse after a few years of hormone therapy. The androgen-independent stage of prostate cancer has been shown to be associated with the development of neuroendocrine differentiation. We previously demonstrated that neuroendocrine prostate cancer cells derived from LNCaP cells overexpress CaV3.2 T-type voltage-dependent calcium channels. We demonstrate here using prostatic acid phosphatase as a marker of prostate secretion and FM1-43 fluorescence imaging of membrane trafficking that neuroendocrine differentiation is associated with an increase in calcium-dependent secretion which critically relies on CaV3.2 T-type calcium channel activity. In addition, we show that these channels are expressed by neuroendocrine cells in prostate cancer tissues obtained from patients after surgery. We propose that CaV3.2 T-type calcium channel up-regulation may account for the alteration of secretion during prostate cancer development and that these channels, by promoting the secretion of potential mitogenic factors, could participate in the progression of the disease toward an androgen-independent stage

Modulation of ER stress and apoptosis by endoplasmic reticulum calcium leak via translocon during unfolded protein response: involvement of GRP78

International audienceThe endoplasmic reticulum (ER) is involved in many cellular functions, including protein folding and Ca2+ homeostasis. The ability of cells to respond to the ER stress is critical for cell survival, and disruption in such regulation can lead to apoptosis. ER stress is accompanied by alterations in Ca2+ homeostasis, and the ER Ca2+ store depletion by itself can induce ER stress and apoptosis. Despite that, the ER Ca2+ leak channels activated in response to the ER stress remain poorly characterized. Here we demonstrate that ER Ca2+ depletion during the ER stress occurs via translocon, the ER protein complex involved in translation. Numerous ER stress inducers stimulate the ER Ca2+ leak that can be prevented by translocon inhibitor, anisomycin. Expression of GRP78, an ER stress marker, increased following treatment with puromycin (a translocon opener) and was suppressed by anisomycin, confirming a primary role of translocon in ER stress induction. Inhibition of ER store depletion by anisomycin significantly reduces apoptosis stimulated by the ER stress inducers. We suggest that translocon opening is physiologically modulated by GRP78, particularly during the ER stress. The ability to modulate the ER Ca2+ permeability and subsequent ER stress can lead to development of a novel therapeutic approach