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. 2018 Dec 17:12:491.
doi: 10.3389/fncel.2018.00491. eCollection 2018.

Astrocyte-Mediated Neuromodulatory Regulation in Preclinical ALS: A Metadata Analysis

Kathleen Jordan  1 , Joseph Murphy  1 , Anjanya Singh  1   2 , Cassie S Mitchell  1
Affiliations

Astrocyte-Mediated Neuromodulatory Regulation in Preclinical ALS: A Metadata Analysis

Kathleen Jordan et al. Front Cell Neurosci. .

Abstract

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by progressive degradation of motoneurons in the central nervous system (CNS). Astrocytes are key regulators for inflammation and neuromodulatory signaling, both of which contribute to ALS. The study goal was to ascertain potential temporal changes in astrocyte-mediated neuromodulatory regulation with transgenic ALS model progression: glutamate, GTL-1, GluR1, GluR2, GABA, ChAT activity, VGF, TNFα, aspartate, and IGF-1. We examine neuromodulatory changes in data aggregates from 42 peer-reviewed studies derived from transgenic ALS mixed cell cultures (neurons + astrocytes). For each corresponding experimental time point, the ratio of transgenic to wild type (WT) was found for each compound. ANOVA and a student's t-test were performed to compare disease stages (early, post-onset, and end stage). Glutamate in transgenic SOD1-G93A mixed cell cultures does not change over time (p > 0.05). GLT-1 levels were found to be decreased 23% over WT but only at end-stage (p < 0.05). Glutamate receptors (GluR1, GluR2) in SOD1-G93A were not substantially different from WT, although SOD1-G93A GluR1 decreased by 21% from post-onset to end-stage (p < 0.05). ChAT activity was insignificantly decreased. VGF is decreased throughout ALS (p < 0.05). Aspartate is elevated by 25% in SOD1-G93A but only during end-stage (p < 0.05). TNFα is increased by a dramatic 362% (p < 0.05). Furthermore, principal component analysis identified TNFα as contributing to 55% of the data variance in the first component. Thus, TNFα, which modulates astrocyte regulation via multiple pathways, could be a strategic treatment target. Overall results suggest changes in neuromodulator levels are subtle in SOD1-G93A ALS mixed cell cultures. If excitotoxicity is present as is often presumed, it could be due to ALS cells being more sensitive to small changes in neuromodulation. Hence, seemingly unsubstantial or oscillatory changes in neuromodulators could wreak havoc in ALS cells, resulting in failed microenvironment homeostasis whereby both hyperexcitability and hypoexcitability can coexist. Future work is needed to examine local, spatiotemporal neuromodulatory homeostasis and assess its functional impact in ALS.

Keywords: ChAT; GABA; GLT-1; TNFα; VGF; aspartate; glutamate.

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Figures

Figure 1
Figure 1
Glutamate, GLT-1, and GluR complex levels in mixed cultures over ALS disease progression. (A) Glutamate level average in SOD1-G93A mice normalized to wild-type (ratio presented as SOD1-G93A/WT) in three temporal stages: 0–96 days, 97–116 days, and 117+ days. There was no significant change in glutamate levels between any of the temporal stages. (B) GLT-1 average in SOD1-G93A normalized to wild-type (ratio presented as SOD1-G93A/WT) in three temporal stages: 0–96 days, 97–116 days, and 117+ days. GLT-1 is significantly decreased from WT (*p = 0.0075) at 117 + days. (C) GluR1 (solid bars) and GluR2 (stripped bars) average in SOD1-G93A normalized to wild-type (ratio presented as SOD1-G93A/WT) in three temporal stages: 0–96 days, 97–116 days, and 117+ days. There was no significant change in GluR2 levels between temporal stages. GluR1 at 117+ days was significantly decreased from 0–96 (*p = 0.0346). Error bars represent SD.
Figure 2
Figure 2
GABA levels over ALS disease progression. GABA level average normalized to wild-type (ratio presented as transgenic/WT) in two temporal stages: 0–96 days and 117+ days. Error bars represent SD.
Figure 3
Figure 3
Calcium-related cytokine levels over ALS disease progression. ChAT activity (A), VGF (B), TNFα (C), and aspartate (D) averages in SOD1-G93A normalized to wild type (ratio presented as SOD1-G93A/WT) in three stages: 0–96 days, 97–116 days, and 117 + days. SOD1-G93A VGF levels were significantly decreased compared to WT at 0–96 days (#p = 0.0036) and 117+ days (#p < 0.0001). SOD1-G93A VGF levels at end-stage are also significantly lower than early-stage (*p = 0.0002). TNFα (*p = 0.0488) and aspartate (#p = 0.0350) levels were significantly increased at end-stage compared to WT. Error bars represent SD.
Figure 4
Figure 4
Temporal relationships between non-inflammatory regulators of astrocytes visualized as a cross-correlation matrix. Average concentrations of glutamate, GABA, and related cytokines for ALS mice were normalized to wild-type mice with data aggregated across three temporal stages: (A) 0–96 days, (B) 97–116 days, and (C) 117+ days. Highly positive correlations were found between VGF and ChAT at 0–96 days. Note that only factors with sufficient sample sizes (calculated using standard statistical power analysis) were included.
Figure 5
Figure 5
PCA to determine variance contribution of non-inflammatory regulators of astrocytes. Biplot showing variable contributions to the first two PCA-determined components (A) broken down to the first two components and the comprising variables (B,C). Component 1 accounted for 55% of the variance and was almost completely comprised of TNFα (B). Component 2 accounted for 17% of the variance and was mostly comprised of GABA and aspartate (C).
Figure 6
Figure 6
Cytokine levels compared to glutamate and GABA levels in mixed cultures over ALS disease progression. VGF (A), TNFα (B), GABA (C), and glutamate (D) averages normalized to wild-type (ratio presented as transgenic/WT) in three temporal stages: 0-96 days, 97-116 days, and 117+ days. ChAT activity (D), aspartate (E), and GABA average normalized to wild-type (ratio presented as transgenic/WT) in two temporal stages: 0–96 days and 117+ days, as insufficient data was available for 97–116 days. VGF levels were significantly decreased from glutamate levels at both stages (*p < 0.05). TNFα levels were significantly increased from glutamate levels at 117+ days (*p = 0.0018). GABA levels were significantly decreased from ChAT levels at 0–96 days (**p = 0.0033) and at 117+ days (**p = 0.009). Error bars represent SD.
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