Effect of β-Hydroxybutyrate on Autophagy Dynamics During Severe Hypoglycemia and the Hypoglycemic Coma.

Carmen Torres-Esquivel,Marco Flores-Méndez,Lourdes Massieu,Teresa Montiel

doi:10.3389/fncel.2020.547215

Carmen Torres-Esquivel, Marco Flores-Méndez + Show 2 more

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https://doi.org/10.3389/fncel.2020.547215

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Abstract

Glucose supply from blood is mandatory for brain functioning and its interruption during acute hypoglycemia or cerebral ischemia leads to brain injury. Alternative substrates to glucose such as the ketone bodies (KB), acetoacetate (AcAc), and β-hydroxybutyrate (BHB), can be used as energy fuels in the brain during hypoglycemia and prevent neuronal death, but the mechanisms involved are still not well understood. During glucose deprivation adaptive cell responses can be activated such as autophagy, a lysosomal-dependent degradation process, to support cell survival. However, impaired or excessive autophagy can lead to cell dysfunction. We have previously shown that impaired autophagy contributes to neuronal death induced by glucose deprivation in cortical neurons and that D isomer of BHB (D-BHB) reestablishes the autophagic flux increasing viability. Here, we aimed to investigate autophagy dynamics in the brain of rats subjected to severe hypoglycemia (SH) without glucose infusion (GI), severe hypoglycemia followed by GI (SH + GI), and a brief period of hypoglycemic coma followed by GI (Coma). The effect of D-BHB administration after the coma was also tested (Coma + BHB). The transformation of LC3-I to LC3-II and the abundance of autophagy proteins, Beclin 1 (BECN1), ATG7, and ATG12–ATG5 conjugate, were analyzed as an index of autophagosome formation, and the levels of sequestrosome1/p62 (SQSTM1/p62) were determined as a hallmark of autophagic degradation. Data suggest that autophagosomes accumulate in the cortex and the hippocampus of rats after SH, likely due to impaired autophagic degradation. In the cortex, autophagosome accumulation persisted at 6 h after GI in animals exposed to SH but recovered basal levels at 24 h, while in the hippocampus no significant effect was observed. In animals subjected to coma, autophagosome accumulation was observed at 24 h after GI in both regions. D-BHB treatment reduced LC3-II and SQSTM1/p62 content and reduced ULK1 phosphorylation by AMPK, suggesting it stimulates the autophagic flux and decreases AMPK activity reducing autophagy initiation. D-BHB also reduced the number of degenerating cells. Together, data suggest different autophagy dynamics after GI in rats subjected to SH or the hypoglycemic coma and support that D-BHB treatment can modulate autophagy dynamics favoring the autophagic flux.

Highlights

The brain is a highly dynamic and energy-demanding organ that depends on the continuous glucose supply from blood, thereby disturbed glucose metabolism can lead to brain dysfunction and even brain injury (Mergenthaler et al, 2013)
During conditions of severe energy failure such as ischemia, hypoxia, and cerebral trauma, neuronal death can result from dysfunctional (Carloni et al, 2008; Sarkar et al, 2014) or excessive autophagy (Shi et al, 2012)
We have previously reported that glucose depletion in cortical cultured neurons activates autophagy, but after glucose reintroduction deficient autophagic degradation is triggered due to calpain-mediated lysosomal dysfunction contributing to neuronal death (Geronimo-Olvera et al, 2017)

Summary

Introduction

The brain is a highly dynamic and energy-demanding organ that depends on the continuous glucose supply from blood, thereby disturbed glucose metabolism can lead to brain dysfunction and even brain injury (Mergenthaler et al, 2013). Reduced cerebral glucose delivery occurs during hypoglycemia, a condition considered as a major complication of insulin treatment in type 1 diabetes mellitus (DMT1) patients (Cryer, 2005). Patients can suffer two events of moderate hypoglycemia (60–40 mg/dl blood glucose) per week and one of severe hypoglycemia (SH, >35 mg/dl) per year. Under conditions of limited glucose availability, such as ischemia, hypoxia, hypoglycemia, and cerebral trauma, alternative energy substrates to glucose, such as the ketone bodies (KB), acetoacetate (AcAc), and β-hydroxybutyrate (BHB), can be used by the brain (Melø et al, 2006) and prevent brain injury (Suzuki et al, 2002; Masuda et al, 2005; Puchowicz et al, 2008; Haces et al, 2008; Julio-Amilpas et al, 2015). We have recently reported that the D isomer of BHB (D-BHB) stimulates the autophagic flux during glucose deprivation and prevents neuronal death in cortical cultures (Camberos-Luna et al, 2016)

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