Intracellular fibril formation, calcification, and enrichment of chaperones, cytoskeletal, and intermediate filament proteins in the adult hippocampus CA1 following neonatal exposure to the nonprotein amino acid BMAA

Abstract

The environmental neurotoxin β-N-methylamino-l-alanine (BMAA) has been implicated in the etiology of neurodegenerative disease, and recent studies indicate that BMAA can be misincorporated into proteins. BMAA is a developmental neurotoxicant that can induce long-term learning and memory deficits, as well as regionally restricted neuronal degeneration and mineralization in the hippocampal CA1. The aim of the study was to characterize long-term changes (2 weeks to 6 months) further in the brain of adult rats treated neonatally (postnatal days 9–10) with BMAA (460 mg/kg) using immunohistochemistry (IHC), transmission electron microscopy, and laser capture microdissection followed by LC-MS/MS for proteomic analysis. The histological examination demonstrated progressive neurodegenerative changes, astrogliosis, microglial activation, and calcification in the hippocampal CA1 3–6 months after exposure. The IHC showed an increased staining for α-synuclein and ubiquitin in the area. The ultrastructural examination revealed intracellular deposition of abundant bundles of closely packed parallel fibrils in neurons, axons, and astrocytes of the CA1. Proteomic analysis of the affected site demonstrated an enrichment of chaperones (e.g., clusterin, GRP-78), cytoskeletal and intermediate filament proteins, and proteins involved in the antioxidant defense system. Several of the most enriched proteins (plectin, glial fibrillar acidic protein, vimentin, Hsp 27, and ubiquitin) are known to form complex astrocytic inclusions, so-called Rosenthal fibers, in the neurodegenerative disorder Alexander disease. In addition, TDP-43 and the negative regulator of autophagy, GLIPR-2, were exclusively detected. The present study demonstrates that neonatal exposure to BMAA may offer a novel model for the study of hippocampal fibril formation in vivo.Electronic supplementary materialThe online version of this article (doi:10.1007/s00204-014-1262-2) contains supplementary material, which is available to authorized users.

Imaging mass spectrometry in drug development and toxicology

“…Ultrastructural examination by transmission electron microscopy revealed intracellular deposition of abundant bundles of closely packed parallel fibrils in neurons, axons and astrocytes of the CA1. SIMS IMS and laser capture microdissection followed by LC–MS/MS showed enrichment of specific phospholipids and proteins in this brain region (Hanrieder et al 2014a, b; Karlsson et al 2015a, b). These molecular studies of BMAA-induced effects further illustrate the usefulness of combining IMS with other experimental techniques to identify target regions and investigate toxicological events.

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β-N-Methylamino-l-alanine (BMAA) perturbs alanine, aspartate and glutamate metabolism pathways in human neuroblastoma cells as determined by metabolic profiling

“…BMAA is considered to have a low neurotoxic potency in adult rodents (Perry et al 1989; Cruz-Aguado et al 2006). However, BMAA is a developmental neurotoxin inducing long-term cognitive deficits as well as neurodegeneration, astrogliosis, and intracellular fibril formation in the hippocampus of adult rodents following neonatal exposure to BMAA (Karlsson et al 2009b, 2014). BMAA also induces behavioral changes in neonates and perturbs serum metabolites that are associated with changes in energy metabolism and amino acid metabolism (Karlsson et al 2009a; Engskog et al 2013).…”

“…Previous studies demonstrated that BMAA is a mixed glutamate receptor agonist that can induce excitotoxicity in primary neurons in vitro and in vivo (Weiss et al 1989; Zeevalk and Nicklas 1989; Chiu et al 2012) but BMAA is also suggested to induce neurotoxicity through other mechanisms such as oxidative stress (Lobner et al 2014). Recent studies have demonstrated enrichment of proteins implicated in protein aggregation, and an increased protein ubiquitination in the adult hippocampus following neonatal administration suggesting BMAA-induced perturbations of cellular protein homeostasis (Karlsson et al 2014). In addition, a dose-dependent increase of protein-associated BMAA in neonatal rat brain, especially in the hippocampus, has been observed (Karlsson et al 2015).…”

MALDI imaging delineates hippocampal glycosphingolipid changes associated with neurotoxin induced proteopathy following neonatal BMAA exposure

“…Exposure to a higher dose (460 mg/kg) also induced altered expression of S100β, histones, calcium and calmodulin-binding proteins as well as guanine nucleotide-binding proteins. At this dose; neuronal degeneration, astrogliosis, intracellular fibril formation, and increased protein ubiquitination were evident in the hippocampus of some of the animals [17,19]. Laser microdissection of tissue samples from the affected hippocampal regions was performed and followed by LC–MS/MS based proteomic analysis.…”