Group I metabotropic receptor antagonism blocks depletion of calcium stores and reduces potentiated capacitative calcium entry in strain-injured neurons and astrocytes

Antagonism of the group I metabotropic receptor subtype 1 (mGluR1) with (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA) has been shown to reduce deficits after in vivo or in vitro traumatic brain injury. We have previously demonstrated that AIDA prevents elevation of astrocyte IP3 subsequent to injury-induced activation of mGluRs and phospholipase C. Since IP3 can cause release of intracellular Ca2+ stores we tested the hypothesis that pre- or post-injury treatment with AIDA can affect (1) the depletion of Ca2+ stores which occurs soon after strain injury of cultured neurons and astrocytes and (2) the delayed potentiation of capacitative calcium entry in strain-injured neurons. Astrocyte or neuronal plus glial cultures were grown on Silastic membranes that were subjected to a 50-msec pulse of compressed gas, which caused membrane displacement and biaxial strain (stretch) injury of the adhering cells. Cells were treated 10 min before or immediately after injury with 100 microM AIDA and the intracellular free Ca2+ ([Ca2+]i) response to thapsigargin, which inhibits the ability of the stores to sequester Ca2+, was measured at 15 min or 3 h after injury. AIDA pre- or post-injury treatment prevented the depletion of intracellular calcium stores at 15 min post-injury in astrocytes and neurons and reduced the potentiated neuronal capacitative calcium influx 3 h after injury. Since Ca2+ and Ca2+ stores influence many factors, including neuronal excitability, plasticity, protein synthesis, and neuronal-glial interactions, prevention of Ca2+ store depletion and subsequent exaggerated capacitative calcium entry may be an important subcellular mechanism by which antagonism of mGluR1 receptors exert an injury-reducing effect. More globally, the results further emphasize the importance of altered signaling and calcium regulatory mechanisms in the immediate and delayed sequelae of traumatic brain injury.     

Store-operated calcium channel inhibition attenuates neutrophil function and postshock acute lung injury

BACKGROUND: A wide variety of neutrophil (PMN) functions are regulated by cytosolic calcium concentration. Calcium channel blockade might therefore decrease postshock inflammation but could also limit important cardiovascular compensations. PMN Ca2+ entry occurs, however, through store-operated calcium entry (SOCE) channels rather than the voltage operated (L-type) channels that regulate cardiovascular tone. We hypothesized that SOCE inhibition might suppress postshock PMN activation, lessening lung injury without compromising cardiovascular performance. METHODS: Human PMNs were treated in vitro with N-propargyl-nitrendipine (MRS1845 [MRS]) a dihydropyridine Ca2+ channel blocker with relative specificity for SOCE channels. Calcium flux was measured by fura fluorescence. Chemotaxis was studied in modified Boyden chambers. Respiratory burst was studied by dihydrorhodamine fluorescence. Exploratory studies were then performed where rats were subjected to trauma and hemorrhagic shock (T/HS) (laparotomy, then hemorrhage to a mean arterial pressure of 30-40 mm Hg for 90 minutes) after pretreatment with MRS or vehicle given intraperitoneally at laparotomy. In vivo PMN CD11b expression was then assayed by flow cytometry and lung injury was assessed as percentage Evans blue dye leak 3 hours after resuscitation. The shed blood volume required to achieve standardized hypotension was measured. RESULTS: In vitro, MRS suppressed human PMN SOCE without affecting calcium store release; it suppressed chemotaxis (60 +/- 6 vs. 150 +/- 15 x 10(3) PMNs/well, p = 0.002) and suppressed respiratory burst (62 +/- 11% vs. 100%, p < 0.05) at IC50 concentrations similar to those needed to suppress SOCE. In subsequent in vivo rat studies, MRS decreased postshock PMN CD11b expression from 397 +/- 93 to 268 +/- 39 MFU mean flourescent units (p < 0.05) and decreased lung Evans blue dye permeability from 8.1 +/- 1.9% to 3.4 +/- 0.1% (p < 0.05). MRS had no noticeable effect on the relationship between blood pressure and blood loss, with shed blood volume remaining almost identical (26 +/- 2 mL/kg vs. 27 +/- 3 mL/kg, p = not significant). CONCLUSION: Modulation of PMN Ca2+ entry by means of selective SOCE channel inhibition attenuates PMN inflammatory responses in vitro. In vivo, SOCE channel blockade attenuates trauma and hemorrhagic shock-induced PMN priming and lung injury without gross evidence of hemodynamic side effects. The relative specificity of SOCE channel blockade for "nonexcitable" cells such as PMNs may make it a valuable form of chemoprophylaxis for the inflammatory consequences of hemorrhagic shock in trauma patients.     

Angiotension converting enzyme inhibition and calcium channel blockage improves cyclosporine induced glucose intolerance in rats

The aim of this study was to evaluate the effects of quinapril, valsartan, and amlodipin on glucose tolerance in cyclosporine (CsA)-toxic rats. Among 40 male Wistar rats 32 were administered cyclosporine (CsA) (15 mg/kg) intraperitoneally for 6 weeks. Quinapril (10 mg/kg per day) (group Q), valsartan (40 mg/kg per day) (group V), and amlodipine (10mg/kg per day) (group A) were administered to individual sets of eight CsA-treated animals via the drinking water with the remaining untreated hosts followed as a control group (group C) and 8 healthy controls (group H). A Glucose-tolerance test was performed by administering oral glucose (2 g/kg) followed by blood samples obtained from the tail vein at baseline as well as 30,60,90, and 120 minutes after the glucose load. Glucose area under the curve (AUC) was calculated according to the trapezoidal rule. CsA levels were determined using an immunofluorescence method. Kruskal Wallis ANOVA test was used for statistical analysis. Median CsA levels were 1968 ng/mL, 1982 ng/mL, 1580 ng/mL, 1600 ng/mL; and glucose AUC, 232.4 +/- 130 mg/min per dL, 63.1 +/- 25 mg/min per dL, 115.0 +/- 90 mg/min per dL, 47.4 +/- 34 mg/min per dL 53.4 +/- 38 mg/min per dL for groups C,Q,V,A and H, respectively. Quinapril-treated and amilodipine-treated rats displayed a lower glucose AUC than group C (P <.01), which had higher glucose levels than healthy controls (P <.001). In summary, CsA treated rats show impaired glucose tolerance, which is improved by quinapril or amlodipine treatment. Angiotensin converting enzymes inhibitors and calcium channel blockers affect beta cell function rather than insulin sensitivity.     

Phaeochromocytoma and calcium channel block

Canadian Journal of Anesthesia/Journal canadien d'anesthésie -     

Irradiation with a low-level diode laser induces the developmental endothelial locus-1 gene and reduces proinflammatory cytokines in epithelial cells

We demonstrated previously that low-level diode laser irradiation with an indocyanine green-loaded nanosphere coated with chitosan (ICG-Nano/c) had an antimicrobial effect, and thus could be used for periodontal antimicrobial photodynamic therapy (aPDT). Since little is known about the effects of aPDT on periodontal tissue, we here investigated the effect of low-level laser irradiation, with and without ICG-Nano/c, on cultured epithelial cells. Human oral epithelial cells were irradiated in a repeated pulse mode (duty cycle, 10 %; pulse width, 100 ms; peak power output, 5 W). The expression of the developmental endothelial locus 1 (Del-1), interleukin-6 (IL-6), IL-8, and the intercellular adhesion molecule-1 (ICAM-1) were evaluated in Ca9-22 cells stimulated by laser irradiation and Escherichia coli-derived lipopolysaccharide (LPS). A wound healing assay was carried out on SCC-25 cells irradiated by diode laser with or without ICG-Nano/c. The mRNA expression of Del-1, which is known to have anti-inflammatory activity, was significantly upregulated by laser irradiation (p?<?0.01). Concurrently, LPS-induced IL-6 and IL-8 expression was significantly suppressed in the LPS?+?laser group (p?<?0.01). ICAM-1 expression was significantly higher in the LPS?+?laser group than in the LPS only or control groups. Finally, compared with the control, the migration of epithelial cells was significantly increased by diode laser irradiation with or without ICG-Nano/c. These results suggest that, in addition to its antimicrobial effect, low-level diode laser irradiation, with or without ICG-Nano/c, can suppress excessive inflammatory responses via a mechanism involving Del-1, and assists in wound healing.     

N-type calcium channel and renal injury

International Urology and Nephrology - Accumulating evidences indicated that voltage-gated calcium channels (VDCC), including L-, T-, N-, and P/Q-type, are present in kidney and contribute to renal...     

Marked disturbance of calcium homeostasis in mice with targeted disruption of the Trpv6 calcium channel gene.

We report the phenotype of mice with targeted disruption of the Trpv6 (Trpv6 KO) epithelial calcium channel. The mice exhibit disordered Ca(2+) homeostasis, including defective intestinal Ca(2+) absorption, increased urinary Ca(2+) excretion, decreased BMD, deficient weight gain, and reduced fertility. Although our Trpv6 KO affects the closely adjacent EphB6 gene, the phenotype reported here is not related to EphB6 dysfunction. INTRODUCTIOn: The mechanisms underlying intestinal Ca(2+) absorption are crucial for overall Ca(2+) homeostasis, because diet is the only source of all new Ca(2+) in the body. Trpv6 encodes a Ca(2+)-permeable cation channel responsible for vitamin D-dependent intestinal Ca(2+) absorption. Trpv6 is expressed in the intestine and also in the skin, placenta, kidney, and exocrine organs. MATERIALS AND METHODS: To determine the in vivo function of TRPV6, we generated mice with targeted disruption of the Trpv6 (Trpv6 KO) gene. RESULTS: Trpv6 KO mice are viable but exhibit disordered Ca(2+) homeostasis, including a 60% decrease in intestinal Ca(2+) absorption, deficient weight gain, decreased BMD, and reduced fertility. When kept on a regular (1% Ca(2+)) diet, Trpv6 KO mice have deficient intestinal Ca(2+) absorption, despite elevated levels of serum PTH (3.8-fold) and 1,25-dihydroxyvitamin D (2.4-fold). They also have decreased urinary osmolality and increased Ca(2+) excretion. Their serum Ca(2+) is normal, but when challenged with a low (0.25%) Ca(2+) diet, Trpv6 KO mice fail to further increase serum PTH and vitamin D, ultimately developing hypocalcemia. Trpv6 KO mice have normal urinary deoxypyridinoline excretion, although exhibiting a 9.3% reduction in femoral mineral density at 2 months of age, which is not restored by treatment for 1 month with a high (2%) Ca(2+) "rescue" diet. In addition to their deranged Ca(2+) homeostasis, the skin of Trpv6 KO mice has fewer and thinner layers of stratum corneum, decreased total Ca(2+) content, and loss of the normal Ca(2+) gradient. Twenty percent of all Trpv6 KO animals develop alopecia and dermatitis. CONCLUSIONS: Trpv6 KO mice exhibit an array of abnormalities in multiple tissues/organs. At least some of these are caused by tissue-specific mechanisms. In addition, the kidneys and bones of Trpv6 KO mice do not respond to their elevated levels of PTH and 1,25-dihydroxyvitamin D. These data indicate that the TRPV6 channel plays an important role in Ca(2+) homeostasis and in other tissues not directly involved in this process.     

Glomerular size-selective dysfunction in NIDDM is not ameliorated by ACE inhibition or by calcium channel blockade

BACKGROUND: In patients with insulin-dependent diabetes mellitus (IDDM) and overt nephropathy glomerular barrier size-selectivity progressively deteriorates with time and is effectively improved by angiotensin converting enzyme (ACE) inhibition. Whether similar glomerular functional changes develop in proteinuric patients with non-insulin-dependent diabetes mellitus (NIDDM), and whether antihypertensive agents can favorably affect glomerular filtration of macromolecules in these patients, has not been documented yet. METHODS: We investigated renal hemodynamics and fractional clearance of neutral dextrans of graded sizes, in nine proteinuric patients with NIDDM and renal biopsy findings of typical diabetic glomerulopathy. Six healthy volunteers served as controls. We also investigated the effects of an ACE inhibitor and of a calcium channel blocker, both given in doses targeted to achieve a comparable level of systemic blood pressure control, on glomerular hemodynamics and sieving function. Theoretical analysis of glomerular macromolecule transport was adopted to evaluate intrinsic glomerular membrane permeability properties. RESULTS: Fractional clearance of large macromolecules (42 to 66 A in radius) was significantly higher in diabetic patients than in controls, and the distribution of membrane pore radii was calculated to be shifted towards larger pore sizes in diabetics (mean radius increased from 55 to 60 A). Despite effective blood pressure control, neither antihypertensive affected glomerular hemodynamics to any significant extent. Fractional clearance of dextrans, as well as of albumin and IgG, and total urinary proteins were not modified by either treatments. CONCLUSIONS: These data indicate that patients with NIDDM and overt nephropathy develop abnormalities in size-selective function of the glomerular barrier and, at variance to IDDM, such changes were not ameliorated either by ACE inhibition or calcium channel blockade.     

T型钙通道与疼痛

T型钙通道具有快速激活,缓慢关闭和较低的电导特性,在静息膜电位下即可激活。T型钙通道广泛分布于中枢和外周神经系统,对调节细胞的兴奋性和神经递质的释放具有重要作用。研究表明,T型钙通道参与了疼痛形成,作用于T型钙通道靶点的镇痛新药具有广阔的前景。     

Epac-Rap signaling reduces cellular stress and ischemia-induced kidney failure

Renal ischemia-reperfusion injury is associated with the loss of tubular epithelial cell-cell and cell-matrix interactions which contribute to renal failure. The Epac-Rap signaling pathway is a potent regulator of cell-cell and cell-matrix adhesion. The cyclic AMP analogue 8-pCPT-2'-O-Me-cAMP has been shown to selectively activate Epac, whereas the addition of an acetoxymethyl (AM) ester to 8-pCPT-2'-O-Me-cAMP enhanced in vitro cellular uptake. Here we demonstrate that pharmacological activation of Epac-Rap signaling using acetoxymethyl-8-pCPT-2'-O-Me-cAMP preserves cell adhesions during hypoxia in vitro, maintaining the barrier function of the epithelial monolayer. Intrarenal administration in vivo of 8-pCPT-2'-O-Me-cAMP also reduced renal failure in a mouse model for ischemia-reperfusion injury. This was accompanied by decreased expression of the tubular cell stress marker clusterin-α, and lateral expression of β-catenin after ischemia indicative of sustained tubular barrier function. Our study emphasizes the undervalued importance of maintaining tubular epithelial cell adhesion in renal ischemia and demonstrates the potential of pharmacological modulation of cell adhesion as a new therapeutic strategy to reduce the extent of injury in kidney disease and transplantation.     

Lack of voltage-dependent calcium channel opening during the calcium influx induced by progesterone in human sperm. Effect of calcium channel deactivation and inactivation

Progesterone induces calcium influx and acrosomal exocytosis in human sperm. Pharmacologic evidence suggests that voltage-dependent calcium channels (VDCCs) are involved. In this study, membrane potential (Vm) and intracellular calcium concentration ([Ca(2+)](i)) were monitored simultaneously to assess the effect of VDCC gating on the calcium influx triggered by progesterone. Holding the Vm to values that maintained VDCCs in a deactivated (-71 mV) closed state inhibited the calcium influx induced by progesterone by approximately 40%. At this Vm, the acrosomal reaction induced by progesterone, but not by A23187, was inhibited. However, when the Vm was held at -15 mV (which maintains VDCCs in an inactivated closed state), the progesterone-induced calcium influx was stimulated. Furthermore, the progesterone and voltage-dependent calcium influxes were additive. These findings indicate that progesterone does not produce VDCC gating in human sperm.