July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Protein expression and functional characterization of muscarinic receptors in myoepithelial cells isolated from rat lacrimal gland
Author Affiliations & Notes
  • Martin Johnsson
    Pharmacology, University of Gothenburg, Gothenburg, Sweden
  • Michael Winder
    Pharmacology, University of Gothenburg, Gothenburg, Sweden
  • Footnotes
    Commercial Relationships   Martin Johnsson, None; Michael Winder, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4164. doi:
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      Martin Johnsson, Michael Winder; Protein expression and functional characterization of muscarinic receptors in myoepithelial cells isolated from rat lacrimal gland. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4164.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Dry eyes disease (DED) is considered a major ocular condition worldwide. DED is more prevalent in the elderly population and ubiquitous modernities such as tablets and smart phones have been shown to increase the incidence of DED. Also, dry eyes is a common adverse effect to many systemic drugs and as such might tamper with patient compliance if not treated. With the multifactorial causality of DED in mind, efforts for pharmacological symptomatic treatment can easily be argued for. In this study the expression of muscarinic receptors and their functional characteristics were investigated in primary myoepithelial cells (MECs). Understanding smooth muscle contractility, and how to modulate it, is of key importance for understanding how lacrimal secretion is regulated.

Methods : The functional response of MECs was investigated by fluorescent spectroscopic quantification of the intracellular calcium indicator FLIPR Calacium 6 (Molecular Devices). Intracellular calcium was quantified as a result of administration of the non-selective muscarinic agonist methacholine in the presence or absence of different muscarinic antagonists with varying muscarinic receptor subtype selectivity, namely 4-DAMP (M1/3/5), methoctramine (M2) and pFHHSiD (M3). Each isolated culture was verified functionally by performing ATP concentration-response experiments, matching previously published data. Expression of muscarinic receptors (M1-M5) in MECs was studied by western blot analysis. Morphological examinations were also performed on each culture isolate by immunocytochemical verification of α-smooth muscle actin expression.

Results : Intracellular calcium investigations showed no correlation between administered muscarinic agonist (10-11- 10-3 M) and increased intracellular concentration of calcium. The absence of calcium response to non-selective muscarinic agonist was additionally observed in the presence of muscarinic antagonists. Western blot analysis revealed MEC expression of muscarinic receptor subtypes M3, M4 and M5.

Conclusions : Muscarinic receptors might play a unique role in MECs, where the signal transduction originating from stimulated muscarinic excitatory receptors might be mediated through other second messengers than calcium.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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