Effect of developmental NMDAR antagonism with CGP 39551 on aspartame-induced hypothalamic and adrenal gene expression

Abstract

RationaleAspartame (L-aspartyl phenylalanine methyl ester) is a non-nutritive sweetener (NNS) approved for use in more than 6000 dietary products and pharmaceuticals consumed by the general public including adults and children, pregnant and nursing mothers. However a recent prospective study reported a doubling of the risk of being overweight amongst 1-year old children whose mothers consumed NNS-sweetened beverages daily during pregnancy. We have previously shown that chronic aspartame (ASP) exposure commencing in utero may detrimentally affect adulthood adiposity status, glucose metabolism and aspects of behavior and spatial cognition, and that this can be modulated by developmental N-methyl-D-aspartate receptor (NMDAR) blockade with the competitive antagonist CGP 39551 (CGP). Since glucose homeostasis and certain aspects of behavior and locomotion are regulated in part by the NMDAR-rich hypothalamus, which is part of the hypothalamic-pituitary-adrenal- (HPA) axis, we have elected to examine changes in hypothalamic and adrenal gene expression in response to ASP exposure in the presence or absence of developmental NMDAR antagonism with CGP, using Affymetrix microarray analysis.ResultsUsing 2-factor ANOVA we identified 189 ASP-responsive differentially expressed genes (DEGs) in the adult male hypothalamus and 2188 in the adrenals, and a further 23 hypothalamic and 232 adrenal genes significantly regulated by developmental treatment with CGP alone. ASP exposure robustly elevated the expression of a network of genes involved in hypothalamic neurosteroidogenesis, together with cell stress and inflammatory genes, consistent with previous reports of aspartame-induced CNS stress and oxidative damage. These genes were not differentially expressed in ASP mice with CGP antagonism. In the adrenal glands of ASP-exposed mice, GABA and Glutamate receptor subunit genes were amongst those most highly upregulated. Developmental NMDAR antagonism alone had less effect on adulthood gene expression and affected mainly hypothalamic neurogenesis and adrenal steroid metabolism. Combined ASP + CGP treatment mainly upregulated genes involved in adrenal drug and cholesterol metabolism.ConclusionASP exposure increased the expression of functional networks of genes involved in hypothalamic neurosteroidogenesis and adrenal catecholamine synthesis, patterns of expression which were not present in ASP-exposed mice with developmental NMDAR antagonism.

Transgenerational transmission of aspartame-induced anxiety and changes in glutamate-GABA signaling and gene expression in the amygdala

“…Therefore, research on aspartame’s neurobehavioral effects has focused on monoamine neurotransmitter signaling. However, there is no consensus in the clinical or preclinical literature regarding aspartame’s effects on brain monoamine content or behaviors such as memory or depression ( 7 , 8 , 13 – 23 ). Similarly, despite reports of aspartame’s carcinogenic ( 24 , 25 ) and metabolic side effects ( 26 – 30 ), a consensus on these topics is lacking.…”

Strain-based and sex-biased differences in adrenal and pancreatic gene expression between KK/HlJ and C57BL/6 J mice

“…Statistical analyses were performed using IBM SPSS statistics software version 20 (SPSS Inc., Chicago, IL) as previously described [27,35]. Data were presented as means ± SEM for body characteristics and Insulin Tolerance test (ITT).…”

“…Total RNA was prepared from snap-frozen male and female adrenal and pancreatic tissue using Qiagen RNeasy Lipid Tissue Mini Kit Cat # 74804 (Qiagen, CA, USA) according to the manufacturer's instructions, and stored at − 80 o C, as described previously [35]. This method was slightly modified for pancreatic RNA extraction, according to De Lisle, 2014 [36].…”

“…Gene expression was analyzed using 12 GeneChip (R) Mouse Gene 2.0 ST arrays representing 26,515 genes as previously described [35]. To minimize the differences of individual variability and increase the statistical power for the identification of potential biomarkers, microarray analysis was performed using equal amounts of purified RNA pooled from all of the study subjects (N = 18 per treatment group), and applied to 3 identical arrays from the same batch.…”

GluN2A/ERK/CREB Signaling Pathway Involved in Electroacupuncture Regulating Hypothalamic-Pituitary-Adrenal Axis Hyperactivity

“…Studies have highlighted the essential roles of NMDAR in brain signal conduction and nerve function ( Yang et al, 2018 ; Zoodsma et al, 2020 ). Previous reports indicated that NMDAR is involved in various stress responses and plays a vital role in coordinating HPA axis activity ( Collison et al, 2018 ; Lin et al, 2019 ). Activation of NMDAR alters synaptic plasticity and evokes calcium spikes in the hypothalamic paraventricular nucleus (PVN) neurons ( Bains and Ferguson, 1999 ).…”