Sex Differences on N-methyl-D-aspartate Receptor Expression along the Rodent Nephron Open Access

Lim, Heeju Jasmine (Spring 2020)

Permanent URL: https://etd.library.emory.edu/concern/etds/xg94hq72j?locale=en
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Abstract

Background: N-methyl-D-aspartate (NMDA) receptors are calcium channels gated by glutamate and glycine. In the kidneys, epithelial NMDA receptors (NMDAr) induce renal vasodilation. Previous data suggest that estrogen supplementation increases renal vasodilation induced by amino acids in aged rats. However, the mechanism of this increase is not known. Additionally, there is no data showing if there is a sex difference in NMDAr expression in kidney. We hypothesized that there is a sex difference in renal NMDAr expression and that it is dependent on Estrogen.

Methods: We evaluated the renal expression of NMDAr type 1 in mouse kidney epithelial cells by immunohistochemistry and immunofluorescence, and in total kidney by Western blot. We also evaluated the expression of NMDAr on a continuous cultured epithelial cell line with characteristics of renal collecting duct principal cells (mpkCCD), with and without the presence of estradiol (50nM) in the media for 24 hours.

Results: NMDAr were found to be present along the nephron, in both cortical and medullar regions, with higher expression on the juxtamedullary and medullary regions. Under confocal microscopy, we confirmed the expression of NMDAr in AQP2 positive cells. Western blot analysis showed an increased expression of NMDAr type 1 in female mouse kidney in comparison with males (3.34±1.6 vs. 0.95±0.3 AU, p=0.01). Twenty-four-hour mpkCCD cell incubation with estrogen increases NMDAr expression (2.3±1.3 vs. 0.9 ±0.58 AU, p=0.05).

Conclusion: There is a sex difference in kidney NMDAr expression which is related to the presence of estrogen. Sex difference in Kidney NMDAr may explain the differences in renal physiologies and disease susceptibilities between males and females.

Table of Contents

Table of Contents:

I.              Chapter 1: Introduction                                                                                            1-2

II.            Chapter 2: Localization NMDAr Along the Nephron                                         

a.    Nephron Anatomy                                                                                              3-4

b.    NMDAr Structure and Function                                                                           5

c.    NMDAr Localization                                                                                            6

d.    Methods: RNA Expression in Micro Dissected Tubule Segments, Mouse Kidney Retrieval, Immunohistochemistry, Immunofluorescence                              6-8

e.    Results: NMDAr RNA Expression, NMDAr Protein Expression, NMDAr presence on CNT and CCD                                                                              8-13

III.          Chapter 3: Sex Differences in NMDAr Expression

a.    Sex Differences in Hypertension                                                                          14

b.    Methods: Immunohistochemistry, Western Blots, Statistics                                15

c.    Results: Difference in NMDAr Expression Between Male and Female Mice 15-17

IV.         Chapter 4: Role of Estrogen in NMDAr Expression

a.    Role of Estrogen in the Kidney                                                                            18

b.    Methods: Estrogen Treatment, Statistics                                                              18

c.    Results: NMDAr Expression in Estrogen-Treated Cells                                      19

V.            Chapter 5: Discussion                                                                                           20-21

VI.          References                                                                                                            23-24

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