Journal: Scientific Reports

Article Title: Ablation of the N-type calcium channel ameliorates diabetic nephropathy with improved glycemic control and reduced blood pressure

doi: 10.1038/srep27192

Figure Lengend Snippet: ( a ) Time course of body weight changes of experimental mice. Body weight in db/db mice significantly increased compared with db /+ mice. ( b ) Kidney weight of experimental mice at 16 weeks of age. db/db Ca v 2.2 −/− mice showed less renal hypertrophy than that of db/db Ca v 2.2 +/+ mice. ( c , d ) Time course of systolic blood pressure changes of db /+ mouse groups ( c ) and db/db mouse groups ( d ). ( e , f ) Urinary catecholamine of experimental mice at 16 weeks of age. db/db Ca v 2.2 −/− mice showed lower levels of urinary noradrenaline ( e ) and adrenaline ( f ) than db/db Ca v 2.2 +/+ mice. db /+ Ca v 2.2 +/+ mice (n = 7, white circles), db /+ Ca v 2.2 +/− mice (n = 6, white triangles), db /+ Ca v 2.2 −/− mice (n = 7, white squares), db/db Ca v 2.2 +/+ mice (n = 8, black circles), db/db Ca v 2.2 +/− mice (n = 8, black triangles), and db/db Ca v 2.2 −/− mice (n = 8, black squares). * p < 0.05, ** p < 0.01 vs. Ca v 2.2 +/+ mice of the same db genotype.

Article Snippet: After blocking, sections were incubated with both goat anti-WT-1 antibody (Santa Cruz Biotechnology, Santa Cruz, CA) and rabbit anti-Ca v 2.2 antibody (Alomone Labs, Jerusalem, Israel) for 1 h at room temperature.

Techniques:

Journal: Scientific Reports

Article Title: Ablation of the N-type calcium channel ameliorates diabetic nephropathy with improved glycemic control and reduced blood pressure

doi: 10.1038/srep27192

Figure Lengend Snippet: ( a ) Time course of 6-h fasting blood glucose concentrations. db/db Ca v 2.2 −/− mice had lower levels of blood glucose than db/db Ca v 2.2 +/+ mice. ( b ) Serum insulin levels at 16 weeks of age. There were no different among db/db mice groups. ( c ) IPGTTs (2 g/kgBW) of db /+ mice at 15 weeks of age. db /+ Ca v 2.2 −/− mice showed better glucose tolerance compared with db /+ Ca v 2.2 +/+ mice. ( d ) IPGTTs (1 g/kgBW) of db/db Ca v 2.2 −/− mice at 15weeks of age. db/db Ca v 2.2 −/− mice exhibited significantly reduced levels of blood glucose compared with db/db Ca v 2.2 +/+ mice. Since the glucometer has a detection limit up to 999 mg/dL, values above the detection limit were treated as 1000 mg/dL. ( e ) Hemoglobin A1c levels at 16 weeks of age. ( f ) Serum insulin levels of IPGTTs. The insulin levels in db/db Ca v 2.2 −/− mice was significantly higher than these of db/db Ca v 2.2 +/+ mice. db /+ Ca v 2.2 +/+ mice (n = 6, white circles), db /+ Ca v 2.2 +/− mice (n = 9, white triangles), db /+ Ca v 2.2 −/− mice (n = 7, white squares), db/db Ca v 2.2 +/+ mice (n = 7, black circles), db/db Ca v 2.2 +/− mice (n = 7, black triangles), and db/db Ca v 2.2 −/− mice (n = 8, black squares) for GTT. ( g , h ) Blood glucose levels in ITTs at 15 weeks of age of db /+ ( g ) and db/db ( h ) mice. db /+ Ca v 2.2 +/+ mice (n = 5, white circles), db /+ Ca v 2.2 +/− mice (n = 9, white triangles), db /+ Ca v 2.2 −/− mice (n = 7, white squares), db/db Ca v 2.2 +/+ mice (n = 9, black circles), db/db Ca v 2.2 +/− mice (n = 9, black triangles), and db/db Ca v 2.2 −/− mice (n = 7, black squares) for ITT. * p < 0.05, ** p < 0.01 compared with Ca v 2.2 +/+ mice of the same db genotype.

Article Snippet: After blocking, sections were incubated with both goat anti-WT-1 antibody (Santa Cruz Biotechnology, Santa Cruz, CA) and rabbit anti-Ca v 2.2 antibody (Alomone Labs, Jerusalem, Israel) for 1 h at room temperature.

Techniques:

Journal: Scientific Reports

Article Title: Ablation of the N-type calcium channel ameliorates diabetic nephropathy with improved glycemic control and reduced blood pressure

doi: 10.1038/srep27192

Figure Lengend Snippet: ( a ) Time course of urinary albumin excretion per milligram creatinine of experimental mice. Urinary albumin excretion of both db/db Ca v 2.2 −/− mice and db/db Ca v 2.2 +/− mice was lower than that of db/db Ca v 2.2 +/+ mice. ( b ) Serum creatinine levels at 16 weeks of age. ( c ) Creatinine clearance at 16 weeks of age. In db/db Ca v 2.2 +/+ mice, creatinine clearance was suppressed to the same level of db/+ mice. ( d ) Immunohistochemical study of Ca v 2.2 in db /+ Ca v 2.2 +/+ mice and db/db Ca v 2.2 +/+ mice. Ca v 2.2 was positive at glomerular cells (arrows). ( e ) Double immunostaining for Ca v 2.2 (brown) and WT1 (blue) shows double positive cells in a glomerulus (insets). db /+ Ca v 2.2 +/− mice (white triangles), db /+ Ca v 2.2 −/− mice (white squares), db/db Ca v 2.2 +/+ mice (black circles), db/db Ca v 2.2 +/− mice (black triangles), and db/db Ca v 2.2 −/− mice (black squares). * p < 0.05, ** p < 0.01 , vs. db/db Ca v 2.2 +/+ mice. Scale bar = 50 μm.

Article Snippet: After blocking, sections were incubated with both goat anti-WT-1 antibody (Santa Cruz Biotechnology, Santa Cruz, CA) and rabbit anti-Ca v 2.2 antibody (Alomone Labs, Jerusalem, Israel) for 1 h at room temperature.

Techniques: Immunohistochemical staining, Double Immunostaining

Journal: Scientific Reports

Article Title: Ablation of the N-type calcium channel ameliorates diabetic nephropathy with improved glycemic control and reduced blood pressure

doi: 10.1038/srep27192

Figure Lengend Snippet: ( a ) Light microscopic analyses were performed at 16 weeks of age, stained with periodic acid-Schiff. db/db Ca v 2.2 −/− mice showed reduced mesangial expansion compared with db/db Ca v 2.2 +/+ mice. Scale bar = 50 μm. ( b ) Mesangial area in a glomerulus at 16 weeks of age. Mesangial area was increased in db/db Ca v 2.2 +/+ mice and was suppressed in db/db Ca v 2.2 −/− mice. ( c ) Immunostaining for nephrin and podocin. db/db Ca v 2.2 −/− mice maintained of nephrin and podocin to the same level with db /+ mice. Scale bar = 50 μm. ( d , e ) Electron microscopic analyses of glomeruli of experimental mice at 16 weeks of age. GBM thickness was ameliorated in db/db Ca v 2.2 −/− mice. Scale bar = 500 nm. ** p < 0.01 , vs. db/db Ca v 2.2 +/+ mice.

Article Snippet: After blocking, sections were incubated with both goat anti-WT-1 antibody (Santa Cruz Biotechnology, Santa Cruz, CA) and rabbit anti-Ca v 2.2 antibody (Alomone Labs, Jerusalem, Israel) for 1 h at room temperature.

Techniques: Staining, Immunostaining

Journal: Scientific Reports

Article Title: Ablation of the N-type calcium channel ameliorates diabetic nephropathy with improved glycemic control and reduced blood pressure

doi: 10.1038/srep27192

Figure Lengend Snippet: ( a ) ERK phosphorylation was increased in a glomerulus, including mesangial cells and podocytes, of db/db Ca v 2.2 +/+ mice. Its increase was ameliorated in a glomerulus of db/db Ca v 2.2 −/− mice. ( b , c ) Mac-2-positive cells in glomeruli increased in db/db Ca v 2.2 +/+ mice compared with those in db /+ Ca v 2.2 +/+ mice and suppressed in db/db Ca v 2.2 −/− mice. ** p < 0.01 . Scale bar = 50 μm.

Article Snippet: After blocking, sections were incubated with both goat anti-WT-1 antibody (Santa Cruz Biotechnology, Santa Cruz, CA) and rabbit anti-Ca v 2.2 antibody (Alomone Labs, Jerusalem, Israel) for 1 h at room temperature.

Techniques:

Journal: Acta Neuropathologica Communications

Article Title: Sleep deprivation of rats increases postsurgical expression and activity of L-type calcium channel in the dorsal root ganglion and slows recovery from postsurgical pain

doi: 10.1186/s40478-019-0868-2

Figure Lengend Snippet: Perioperative SD increases expression of L-type HVA channels in lumbar (L4–6) DRG neurons, mainly in medium/large cells. a Representative immunofluorescence results, showing higher expression of L-type channels on the neuronal membranes of rats in the incision+SD group than the incision group. Scale bar: 100 μm. b Quantitation of the results in A (10 rats per group). One-way ANOVA followed by a post hoc Tukey test: F(3, 8) = 74.13, ** p < 0.01 for the incision group vs. the incision+SD group. c Size distribution of L-type-channel (Ca v 1.2)-labelled neuronal somata in the DRG (small: 6.59%, medium: 42.86%, large: 50.55%). d Representative immunofluorescence results showing strong co-expression of the L-type channel (Ca v 1.2) with neurofilament 200 (top row), but little or no co-expression with calcitonin gene-related peptide (CGRP; middle row), or isolectin B4 (IB4; bottom row). Arrows indicate co-labeling. Co-labeling analysis used the colocalization finder in Image J. Pearson’s correlation coefficients (r) are indicated. Scale bar: 50 μm

Article Snippet: The membranes were blocked with 1% bovine serum albumin (BSA) at 4 °C overnight, and then incubated with rabbit anti-Ca v 1.2 antibody (L-type; 1:200, ACC-003, Alomone), rabbit anti-Ca v 2.1 antibody (P/Q-type; 1:200, ACC-001, Alomone), rabbit anti-Ca v 2.2 antibody (N-type; 1:200, ACC-002, Alomone), rabbit anti-Ca v 2.3 antibody (R-type; 1:200, ACC-006, Alomone), rabbit anti-Ca v 3.2 antibody (T-type; 1:200, ACC-001, Alomone), mouse anti-Egr1 antibody(1:200, sc-101,033, Santa Cruz), or rabbit beta tubulin antibody (1:3000, AB0039, Abways) at 4 °C overnight under gentle agitation.

Techniques: Expressing, Immunofluorescence, Quantitation Assay, Labeling

Journal: Journal of Ovarian Research

Article Title: Expression of voltage-gated Ca 2+ channels, Insp 3 Rs, and RyRs in the immature mouse ovary

doi: 10.1186/s13048-022-01015-y

Figure Lengend Snippet: Quantification of specific immunolabelling of voltage-gated and intracellular release Ca 2+ channels in the neonatal (left column), early infantile (middle column), and late infantile (right column) ovary. The fluorescence intensity of voltage-gated Ca V 1.2 ( A , A 1 , A 2 ), Ca V 1.3 ( B , B 1 , B 2 ), Ca V 2.1 ( C , C 1 , C 2 ), and Ca V 2.2 ( D, D 1 , D 2 ), as well as InsP 3 R ( E , E 1 , E 2 ) and RyR ( F , F 1 , F 2 ) from the different ovarian structures (primordial, primary, and secondary oocytes, granulosa cells, theca cells, and interfollicular stroma), was measured. The mean fluorescence intensities (arbitrary units) were plotted as bar graphs. A-F : PND3. A 1 -F 1 : PND8. A 2 -F 2 : PND16. The standard error of the mean of each bar is indicated and the number of objects measured of each type. The differences between bars are statistically significant ( p <0.05), except for a few bars in Figures A1, D1, and F1

Article Snippet: Then, sections were incubated for 24 hrs. in a humid chamber at 4°C with one of the primary rabbit antibodies from Alomone Labs (Jerusalem, Israel): anti-Ca V 2.1 ( α 1A; validation number 2039764, lot # AN-09), anti-Ca V 2.2 ( α 1B; validation number 2039766, lot # AN-015), anti-Ca V 1.2 ( α 1C; validation number 2039771, lot # AN-19) or anti-Ca V 1.3 ( α 1D; validation number 2039775, lot # AN-09 : dilution 1:100), rabbit anti-InsP 3 R (I, II, III) H-300 Santa Cruz Biotechnologies (sc-28613; dilution 1:50) and mouse anti-RyR C3-33, (ab2827; dilution 1:20) from ABCAM.

Techniques: Fluorescence

Journal: Journal of Ovarian Research

Article Title: Expression of voltage-gated Ca 2+ channels, Insp 3 Rs, and RyRs in the immature mouse ovary

doi: 10.1186/s13048-022-01015-y

Figure Lengend Snippet: Tissue distribution of specific immunolabelling for Ca V 2.1 and Ca V 2.2 voltage-gated Ca 2+ channels in the neonatal ovary (PND 3). A: Ca V 2.1 specific immunostaining. a , b: Enlarged images from the square areas indicated in A . yellow and red asterisks: Strong staining of primordial and primary follicles, respectively. yellow arrows: Ca V 2.1 positive granulosa cells surrounding primary follicles. blue arrows: Intensively Ca V 2.1 immunoreactive cells (possibly autonomic neurons). blue asterisks: unstained stromal cells. B: Ca V 2.2 specific immunostaining. c , d: Enlarged images from the square areas indicated in B . yellow and red asterisks: Strong Ca V 2.2 immunostaining of primordial and primary follicles, respectively. yellow arrows: positive granulosa cells surrounding primary follicles blue asterisks: unstained stromal cells. Calibration bar: 100 μm (A, B); 50 μm (a, b, c, d)

Article Snippet: Then, sections were incubated for 24 hrs. in a humid chamber at 4°C with one of the primary rabbit antibodies from Alomone Labs (Jerusalem, Israel): anti-Ca V 2.1 ( α 1A; validation number 2039764, lot # AN-09), anti-Ca V 2.2 ( α 1B; validation number 2039766, lot # AN-015), anti-Ca V 1.2 ( α 1C; validation number 2039771, lot # AN-19) or anti-Ca V 1.3 ( α 1D; validation number 2039775, lot # AN-09 : dilution 1:100), rabbit anti-InsP 3 R (I, II, III) H-300 Santa Cruz Biotechnologies (sc-28613; dilution 1:50) and mouse anti-RyR C3-33, (ab2827; dilution 1:20) from ABCAM.

Techniques: Immunostaining, Staining

Journal: Journal of Ovarian Research

Article Title: Expression of voltage-gated Ca 2+ channels, Insp 3 Rs, and RyRs in the immature mouse ovary

doi: 10.1186/s13048-022-01015-y

Figure Lengend Snippet: Tissue distribution of specific immunolabelling for Ca V 2.1 and Ca V 2.2 voltage-gated Ca 2+ channels in the early infantile ovary (PND 8) . A: Ca V 2.1 specific immunostaining. a , b: Enlarged images from the square areas indicated in A . yellow asterisks: weak staining of few primordial oocytes near the plasma membrane. red asterisks: distinct Ca V 2.1 staining of oocytes and granulosa cells from primary and secondary follicles. blue arrows: cytoplasmic staining of granulosa cells close to the plasma membrane in primary follicles. yellow arrows: flat, perifollicular stromal cells next to secondary and early antral follicles show the strongest Ca V 2.1 immunolabeling. B: Ca V 2.2 specific immunostaining. c , d: Enlarged images from the square areas indicated in B . Ca V 2.2-specific staining is moderate throughout the ovary. yellow asterisks: few remaining primordial follicles . red asterisks: oocytes showing moderate Ca V 2.2 immunostaining: blue arrows: weakly stained granulosa cells surrounding oocytes from primary follicles. yellow arrows: patches of perifollicular cells around secondary and early antral follicles show the strongest Ca V 2.2 labeling. Calibration bar: 100 μm (A, B); 50 μm (a, b, c, d)

Article Snippet: Then, sections were incubated for 24 hrs. in a humid chamber at 4°C with one of the primary rabbit antibodies from Alomone Labs (Jerusalem, Israel): anti-Ca V 2.1 ( α 1A; validation number 2039764, lot # AN-09), anti-Ca V 2.2 ( α 1B; validation number 2039766, lot # AN-015), anti-Ca V 1.2 ( α 1C; validation number 2039771, lot # AN-19) or anti-Ca V 1.3 ( α 1D; validation number 2039775, lot # AN-09 : dilution 1:100), rabbit anti-InsP 3 R (I, II, III) H-300 Santa Cruz Biotechnologies (sc-28613; dilution 1:50) and mouse anti-RyR C3-33, (ab2827; dilution 1:20) from ABCAM.

Techniques: Immunostaining, Staining, Immunolabeling, Labeling

Journal: Journal of Ovarian Research

Article Title: Expression of voltage-gated Ca 2+ channels, Insp 3 Rs, and RyRs in the immature mouse ovary

doi: 10.1186/s13048-022-01015-y

Figure Lengend Snippet: Tissue distribution of specific immunolabelling for Ca V 2.1 and Ca V 2.2 voltage-gated Ca 2+ channels in the late infantile ovary (PND 16). A: Ca V 2.1 specific immunostaining. a , b: Enlarged images from the square areas indicated in A. red asterisks: oocytes and granulosa cells from primary, secondary, pre-antral, and antral follicles are express Ca V 2.1 weakly. blue arrows: in contrast, patches of perifollicular cells forming incomplete envelopes around secondary, early antral, and antral follicles are strongly stained. B: Ca V 2.2 specific immunostaining. c , d: Enlarged images from the square areas indicated in B. Weak to moderate Ca V 2.2 labeling is visible throughout the ovary. red asterisks: weakly stained granulosa cells. yellow arrows: most oocytes are weakly stained. green arrows : some oocytes display patches of intense labeling. blue arrows: patches of strongly Ca V 2.2 positive perifollicular cells form an incomplete envelope around early antral and antral follicles. yellow asterisks strongly Ca V 2.2 positive bundles of smooth muscle cells seen at the hilum. Calibration bar: 100 μm (A, B); 50 μm (a, b, c, d)

Article Snippet: Then, sections were incubated for 24 hrs. in a humid chamber at 4°C with one of the primary rabbit antibodies from Alomone Labs (Jerusalem, Israel): anti-Ca V 2.1 ( α 1A; validation number 2039764, lot # AN-09), anti-Ca V 2.2 ( α 1B; validation number 2039766, lot # AN-015), anti-Ca V 1.2 ( α 1C; validation number 2039771, lot # AN-19) or anti-Ca V 1.3 ( α 1D; validation number 2039775, lot # AN-09 : dilution 1:100), rabbit anti-InsP 3 R (I, II, III) H-300 Santa Cruz Biotechnologies (sc-28613; dilution 1:50) and mouse anti-RyR C3-33, (ab2827; dilution 1:20) from ABCAM.

Techniques: Immunostaining, Staining, Labeling