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Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells

PURPOSE: The functional characteristics of receptors that regulate lacrimal gland myoepithelial cells are still somewhat unclear. To date, mainly muscarinic receptors have been of interest; however, further knowledge is needed regarding their expression and functional roles. For this purpose, primar...

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Autores principales: Dankis, Martin, Carlsson, Thomas, Aronsson, Patrik, Tobin, Gunnar, Winder, Michael
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Association for Research in Vision and Ophthalmology 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8458779/
https://www.ncbi.nlm.nih.gov/pubmed/34546325
http://dx.doi.org/10.1167/iovs.62.12.19
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author Dankis, Martin
Carlsson, Thomas
Aronsson, Patrik
Tobin, Gunnar
Winder, Michael
author_facet Dankis, Martin
Carlsson, Thomas
Aronsson, Patrik
Tobin, Gunnar
Winder, Michael
author_sort Dankis, Martin
collection PubMed
description PURPOSE: The functional characteristics of receptors that regulate lacrimal gland myoepithelial cells are still somewhat unclear. To date, mainly muscarinic receptors have been of interest; however, further knowledge is needed regarding their expression and functional roles. For this purpose, primary cultures of rat lacrimal gland myoepithelial cells were established and examined functionally. METHODS: Rat lacrimal glands were excised, minced, and further digested, yielding mixtures of cells that were seeded in culturing flasks. After 4-6 weeks, primary monocultures of myoepithelial cells were established, verified by immunocytochemistry. The cells were stained for all muscarinic receptor subtypes (M1–M5) and examined functionally regarding intracellular [Ca(2+)] responses upon activation of muscarinic receptors. For methodological verification, purinergic functional responses were also studied. RESULTS: Expression of muscarinic receptor subtypes M2-M5 was detected, whereas expression of muscarinic M1 receptors could not be shown. Activation of muscarinic receptors by the non-selective muscarinic agonist methacholine (3 × 10(−11)–10(−3) M) did not cause a significant increase in intracellular [Ca(2+)]. However, activation of purinergic receptors by the non-selective purinergic agonist ATP (10(−8)–10(−3) M) caused a concentration-dependent increase in intracellular [Ca(2+)] that could be blocked by the P2 antagonists PPADS and suramin. CONCLUSIONS: Primary cultures of rat lacrimal gland myoepithelial cells were established that displayed a heterogeneous expression of muscarinic receptors. Purinergic functional responses demonstrated a viable cell population. Upon treatment with methacholine, no significant increase in intracellular [Ca(2+)] could be detected, indicating that cholinergic activation of myoepithelial cells occurs via other intracellular messengers or is dependent on interaction with other cell types.
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spelling pubmed-84587792021-10-05 Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells Dankis, Martin Carlsson, Thomas Aronsson, Patrik Tobin, Gunnar Winder, Michael Invest Ophthalmol Vis Sci Biochemistry and Molecular Biology PURPOSE: The functional characteristics of receptors that regulate lacrimal gland myoepithelial cells are still somewhat unclear. To date, mainly muscarinic receptors have been of interest; however, further knowledge is needed regarding their expression and functional roles. For this purpose, primary cultures of rat lacrimal gland myoepithelial cells were established and examined functionally. METHODS: Rat lacrimal glands were excised, minced, and further digested, yielding mixtures of cells that were seeded in culturing flasks. After 4-6 weeks, primary monocultures of myoepithelial cells were established, verified by immunocytochemistry. The cells were stained for all muscarinic receptor subtypes (M1–M5) and examined functionally regarding intracellular [Ca(2+)] responses upon activation of muscarinic receptors. For methodological verification, purinergic functional responses were also studied. RESULTS: Expression of muscarinic receptor subtypes M2-M5 was detected, whereas expression of muscarinic M1 receptors could not be shown. Activation of muscarinic receptors by the non-selective muscarinic agonist methacholine (3 × 10(−11)–10(−3) M) did not cause a significant increase in intracellular [Ca(2+)]. However, activation of purinergic receptors by the non-selective purinergic agonist ATP (10(−8)–10(−3) M) caused a concentration-dependent increase in intracellular [Ca(2+)] that could be blocked by the P2 antagonists PPADS and suramin. CONCLUSIONS: Primary cultures of rat lacrimal gland myoepithelial cells were established that displayed a heterogeneous expression of muscarinic receptors. Purinergic functional responses demonstrated a viable cell population. Upon treatment with methacholine, no significant increase in intracellular [Ca(2+)] could be detected, indicating that cholinergic activation of myoepithelial cells occurs via other intracellular messengers or is dependent on interaction with other cell types. The Association for Research in Vision and Ophthalmology 2021-09-21 /pmc/articles/PMC8458779/ /pubmed/34546325 http://dx.doi.org/10.1167/iovs.62.12.19 Text en Copyright 2021 The Authors https://creativecommons.org/licenses/by/4.0/This work is licensed under a Creative Commons Attribution 4.0 International License.
spellingShingle Biochemistry and Molecular Biology
Dankis, Martin
Carlsson, Thomas
Aronsson, Patrik
Tobin, Gunnar
Winder, Michael
Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title_full Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title_fullStr Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title_full_unstemmed Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title_short Novel Insights Into Muscarinic and Purinergic Responses in Primary Cultures of Rat Lacrimal Gland Myoepithelial Cells
title_sort novel insights into muscarinic and purinergic responses in primary cultures of rat lacrimal gland myoepithelial cells
topic Biochemistry and Molecular Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8458779/
https://www.ncbi.nlm.nih.gov/pubmed/34546325
http://dx.doi.org/10.1167/iovs.62.12.19
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