isolation-enteric-glial-cells-from-submucosa-lamina-propria-adult

Isolation of Enteric Glial Cells from the Submucosa and Lamina Propria of the Adult Mouse

The enteric nervous system (ENS) consists of neurons and enteric glial cells (EGCs) that reside within the smooth muscle wall, submucosa and lamina ...

The integrity of the plasma membrane is maintained through an active repair process, especially in skeletal and cardiac muscle cells, in which contraction-induced mechanical damage frequently occurs in vivo. Muscular dystrophies (MDs) are a group of muscle diseases characterized by skeletal muscle wasting and weakness. An important cause of these group of diseases is defective repair of sarcolemmal injuries, which normally requires Ca(2+) sensor proteins and Ca(2+)-dependent delivery of intracellular vesicles to the sites of injury. MCOLN1 (also known as TRPML1, ML1) is an endosomal and lysosomal Ca(2+) channel whose human mutations cause mucolipidosis IV (ML4), a neurodegenerative disease with motor disabilities. Here we report that ML1-null mice develop a primary, early-onset MD independent of neural degeneration. Although the dystrophin-glycoprotein complex and the known membrane repair proteins are expressed normally, membrane resealing was defective in ML1-null muscle fibers and also upon acute and pharmacological inhibition of ML1 channel activity or vesicular Ca(2+) release. Injury facilitated the trafficking and exocytosis of vesicles by upmodulating ML1 channel activity. In the dystrophic mdx mouse model, overexpression of ML1 decreased muscle pathology. Collectively, our data have identified an intracellular Ca(2+) channel that regulates membrane repair in skeletal muscle via Ca(2+)-dependent vesicle exocytosis.

The intracellular Ca²⁺ channel MCOLN1 is required for sarcolemma repair to prevent muscular dystrophy.

The resting membrane potential (Δψ) of the cell is negative on the cytosolic side and determined primarily by the plasma membrane's selective permeability to K We show that lysosomal Δψ is set by lysosomal membrane permeabilities to Na and H, but not K, and is positive on the cytosolic side. An increase in juxta-lysosomal Ca rapidly reversed lysosomal Δψ by activating a large voltage-dependent and K-selective conductance (LysoK). LysoK is encoded molecularly by SLO1 proteins known for forming plasma membrane BK channels. Opening of single LysoK channels is sufficient to cause the rapid, striking changes in lysosomal Δψ. Lysosomal Ca stores may be refilled from endoplasmic reticulum (ER) Ca via ER-lysosome membrane contact sites. We propose that LysoK serves as the perilysosomal Ca effector to prime lysosomes for the refilling process. Consistently, genetic ablation or pharmacological inhibition of LysoK, or abolition of its Ca sensitivity, blocks refilling and maintenance of lysosomal Ca stores, resulting in lysosomal cholesterol accumulation and a lysosome storage phenotype.

A voltage-dependent K channel in the lysosome is required for refilling lysosomal Ca stores.

Gastric acid secretion by parietal cells requires trafficking and exocytosis of H/K-ATPase-rich tubulovesicles (TVs) toward apical membranes in response to histamine stimulation via cyclic AMP elevation. Here, we found that TRPML1 (ML1), a protein that is mutated in type IV mucolipidosis (ML-IV), is a tubulovesicular channel essential for TV exocytosis and acid secretion. Whereas ML-IV patients are reportedly achlorhydric, transgenic overexpression of ML1 in mouse parietal cells induced constitutive acid secretion. Gastric acid secretion was blocked and stimulated by ML1 inhibitors and agonists, respectively. Organelle-targeted Ca imaging and direct patch-clamping of apical vacuolar membranes revealed that ML1 mediates a PKA-activated conductance on TV membranes that is required for histamine-induced Ca release from TV stores. Hence, we demonstrated that ML1, acting as a Ca channel in TVs, links transmitter-initiated cyclic nucleotide signaling with Ca-dependent TV exocytosis in parietal cells, providing a regulatory mechanism that could be targeted to manage acid-related gastric diseases.

Gastric Acid Secretion from Parietal Cells Is Mediated by a Ca Efflux Channel in the Tubulovesicle.

The multiple endocrine neoplasia, type 1 (MEN1) locus encodes the nuclear protein and tumor suppressor menin. MEN1 mutations frequently cause neuroendocrine tumors such as gastrinomas, characterized by their predominant duodenal location and local metastasis at time of diagnosis. Diffuse gastrin cell hyperplasia precedes the appearance of MEN1 gastrinomas, which develop within submucosal Brunner's glands. We investigated how menin regulates expression of the gastrin gene and induces generation of submucosal gastrin-expressing cell hyperplasia.

Article	Year
The intracellular Ca²⁺ channel MCOLN1 is required for sarcolemma repair to prevent muscular dystrophy. Nature medicine\| PubMed ID: 25216637	2014
A voltage-dependent K channel in the lysosome is required for refilling lysosomal Ca stores. The Journal of cell biology\| PubMed ID: 28468834	2017
Gastric Acid Secretion from Parietal Cells Is Mediated by a Ca Efflux Channel in the Tubulovesicle. Developmental cell\| PubMed ID: 28486130	2017
Gastrin Induces Nuclear Export and Proteasome Degradation of Menin in Enteric Glial Cells. Gastroenterology\| PubMed ID: 28859856	2017

Nirakar Sahoo

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