Endothelin-Induced Sarcoplasmic Reticulum Calcium Depletion Waves in Vascular Smooth Muscle Cells

Agonist-stimulated waves of elevated cytoplasmic Ca2+ concentration ([Ca2+]i ) regulate blood vessel tone and vasomotion in vascular smooth muscle. Previous studies employing cytoplasmic Ca2+ indicators revealed that these Ca2+ waves were generated by a combination of inositol 1,4,5-trisphosphate (IP3) and Ca2+ induced Ca2+ release (CICR) from the sarcoplasmic reticulum (SR); although, some of the mechanistic details remain uncertain. However, these findings were derived indirectly from observing agonist-induced [Ca2+]i fluctuations in the cytoplasm.
Here, for the first time, we have recorded Endothelin-1 (ET-1) induced waves of Ca2+ depletion from the SR lumen in vascular smooth muscle cells (VSMCs) using a calsequestrin-targeted Ca2+ indicator. Our findings show that these waves: (1) are due to regenerative CICR by the receptors for IP3 (IP3R), (2) have a marked latency period, (3) are characterized by a transient increase in SR Ca2+ ([Ca2+]SR ) both at the point of origin and at the wave front, (4) proceed with diminishing velocity, and (5) are arrested by the nuclear envelope. Our quantitative model indicates that the gradual decrease in the velocity of the SR depletion wave, in the absence of external Ca2+, results from continuity of the SR luminal network

High resolution structural evidence suggests the Sarcoplasmic Reticulum forms microdomains with acidic stores (lysosomes) in the heart

Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) stimulates calcium release from acidic stores such as lysosomes and is a highly potent calcium-mobilising second messenger. NAADP plays an important role in calcium signalling in the heart under basal conditions and following β-adrenergic stress. Nevertheless, the spatial interaction of acidic stores with other parts of the calcium signalling apparatus in cardiac myocytes is unknown. We present evidence that lysosomes are intimately associated with the sarcoplasmic reticulum (SR) in ventricular myocytes; a median separation of 20 nm in 2D electron microscopy and 3.3 nm in 3D electron tomography indicates a genuine signalling microdomain between these organelles. Fourier analysis of immunolabelled lysosomes suggests a sarcomeric pattern (dominant wavelength 1.80 μm). Furthermore, we show that lysosomes form close associations with mitochondria (median separation 6.2 nm in 3D studies) which may provide a basis for the recently-discovered role of NAADP in reperfusion-induced cell death. The trigger hypothesis for NAADP action proposes that calcium release from acidic stores subsequently acts to enhance calcium release from the SR. This work provides structural evidence in cardiac myocytes to indicate the formation of microdomains between acidic and SR calcium stores, supporting emerging interpretations of NAADP physiology and pharmacology in heart

The effect of topical bromfenac on intraretinal and subretinal fluid in neovascular age-related macular degeneration

Purpose: To report the effect of topical bromfenac, a non-steroidal anti-inflammatory drug (NSAID), in a case of neovascular age-related macular degeneration (AMD). Methods: An 85-year-old woman presented with a complaint of visual acuity reduction in the right eye. Comprehensive ophthalmological examination and retinal imaging were performed. Results: Best corrected visual acuity was 2/100. Fundus examination showed reticular pseudodrusen and a small hemorrhage in the fovea. Fluorescein angiography showed an active neovascular membrane. Spectral-domain optical coherence tomography (SD-OCT) confirmed diagnosis and revealed subretinal and intraretinal fluid. The patient refused recommended intravitreal anti-vascular endothelial growth factor treatment and received topical bromfenac 0.09% twice daily. Follow-up with SD-OCT showed subretinal followed by intraretinal fluid reduction at 16 weeks after treatment. Conclusion: Short-term reduction of subretinal and intraretinal fluid was observed with topical bromfenac monotherapy in neovascular AMD