New research sheds light on the underlying mechanisms of 2,4 DABA neurotoxic effects

A new paper by S. Spacic et al. in Aquatic Toxicology, presents important findings with regards to the mechanisms underlying 2,4-DABA neurotoxicity. From the abstract:

“Recent studies suggest that 2,4-DABA, a neurotoxic excitatory amino acid present in virtually all environments, but predominantly in aquatic ecosystems may be a risk factor for development of neurodegenerative diseases in animals and humans. Despite its neurotoxicity and potential environmental importance, mechanisms underlying the excitatory and putative excitotoxic action of 2,4-DABA in neurons are still unexplored. We previously reported on extensive two-stage membrane depolarization and functional disturbances in leech Retzius neurons induced by 2,4-DABA. Current study presents the first detailed look into the electrophysiological processes leading to this depolarization. Intracellular recordings were performed on Retzius neurons of the leech Haemopis sanguisuga using glass microelectrodes and input membrane resistance (IMR) was measured by injecting hyperpolarizing current pulses through these electrodes. Results show that the excitatory effect 2,4-DABA elicits on neurons’ membrane potential is dependent on sodium ions. Depolarizing effect of 5·10−3 mol/L 2,4-DABA in sodium-free solution was significantly diminished by 91% reducing it to 3.26 ± 0.62 mV and its two-stage nature was abrogated. In addition to being sodium-dependent, the depolarization of membrane potential induced by this amino acid is coupled with an increase of membrane permeability, as 2,4-DABA decreases IMR by 8.27 ± 1.47 MΩ (67.60%). Since present results highlight the role of sodium ions, we investigated the role of two putative sodium-dependent mechanisms in 2,4-DABA-induced excitatory effect – activation of ionotropic glutamate receptors and the electrogenic transporter for neutral amino acids. Excitatory effect of 5·10−3 mol/L 2,4-DABA was partially blocked by 10-5 mol/L 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) a non-NMDA receptor antagonist as the first stage of membrane depolarization was significantly reduced by 2.59 ± 0.98 mV (40%), whilst second stage remained unaltered. Moreover, involvement of the sodium-dependent transport system for neutral amino acids was investigated by equimolar co-application of 5·10−3 mol/L 2,4-DABA and L-alanine, a competitive inhibitor of this transporter. Although L-alanine exhibited no effect on the first stage of membrane depolarization elicited by 2,4-DABA, it substantially reduced the second stage (the overall membrane depolarization) from 39.63 ± 2.22 mV to 16.28 ± 2.58 mV, by 58.92%. We therefore propose that the electrophysiological effect of 2,4-DABA on Retzius neurons is mediated by two distinct mechanisms, i.e. by activation of ionotropic glutamate receptor that initiates the first stage of membrane depolarization followed by the stimulation of an electrogenic sodium-dependent neutral amino acid transporter, leading to additional influx of positive charge into the cell and the second stage of depolarization.”

The paper acknowledges CYANOCOST.

Reference:

Svetolik Spasic, Marija Stanojevic, Jelena Nesovic Ostojic, Sanjin Kovacevic, Jasna Todorovic, Marko Dincic, Vladimir Nedeljkov, Milica Prostran, Srdjan Lopicic (2020).
Two distinct electrophysiological mechanisms underlie extensive depolarization elicited by 2,4 diaminobutyric acid in leech Retzius neurons. Aquatic Toxicology 220, 105398.
https://doi.org/10.1016/j.aquatox.2019.105398.

 

New research shows that 2,4-DABA induces irreversible functional changes in neurons.

From the abstract of a recent paper by Spacic et al. in Aquatic Toxicology:

“In this paper we present, for the first time, a detailed account of electrophysiological effects of 2,4-diaminobutyric acid (2,4-DABA). 2,4-DABA is a neurotoxic non-protein amino acid produced by Cyanobacteria with a possible link to neurodegenerative disorders in animals and humans. Intracellular recordings were performed on Retzius nerve cells of the leech Haemopis sanguisuga using glass microelectrodes filled with 3 mol/L KCl. Our results show that 2,4-DABA is an excitatory amino acid, causing membrane depolarization in a concentration dependent manner. The most prominent depolarizations of 39.63 ± 2.22 mV and 47.05 ± 4.33 mV, induced by 5 × 10−3 and 10−2 mol/L 2,4-DABA respectively, are several times larger than maximal depolarizations induced by either Glutamate, Aspartate, β-N-methylamino-alanine (BMAA) or β-N-oxalylamino-alanine (BOAA) on our model. These 2,4-DABA induced depolarizations evolve through two distinct stages, which is a novel phenomenon in electrical cell activity upon application of an excitatory amino acid, at least on our model. Involvement of two separate mechanisms, suggested by the two stage phenomenon, is discussed in the paper. We also provide evidence that 2,4-DABA induces irreversible functional disturbances in neurons in a concentration dependent manner, since only half of the cells recovered normal electrical activity after application of 5 × 10−3 mol/L 2,4-DABA, and none recovered after application of 10−2 mol/L 2,4-DABA. Effects of both L-2,4-DABA and DL-2,4-DABA were tested and are not significantly different.”

Reference:

S. Spasic, M. Stanojevic, J. Nesovic Ostojic, S. Kovacevic, M. Prostran, S. Lopicic (2018).
Extensive depolarization and lack of recovery of leech Retzius neurons caused by 2,4 diaminobutyric acid. Aquatic Toxicology, Volume 199, Pages 269-275. https://doi.org/10.1016/j.aquatox.2018.03.036.

The paper acknowledges CYANOCOST.

Repeated five-day administration of L-BMAA, microcystin-LR, or as mixture, in adult C57BL/6 mice – lack of adverse cognitive effects

Abstract from the Myhre et al. (2018) paper in Scientific Reports:

“The cyanobacterial toxins β-methylamino-L-alanine (L-BMAA) and microcystin-LR (MC-LR; a potent liver toxin) are suspected to cause neurological disorders. Adult male C57BL/6JOlaHsd mice aged approximately 11 months were subcutaneously injected for five consecutive days with L-BMAA and microcystin-LR alone, or as a mixture. A dose-range study determined a tolerable daily dose to be ~31 µg MC-LR/kg BW/day based on survival, serum liver status enzymes, and relative liver and kidney weight. Mice tolerating the first one-two doses also tolerated the subsequent three-four doses indicating adaptation. The LD50 was 43–50 μg MC-LR/kg BW. Long-term effects (up to 10 weeks) on spatial learning and memory performance was investigated using a Barnes maze, were mice were given 30 µg MC-LR/kg BW and/or 30 mg L-BMAA/kg BW either alone or in mixture for five consecutive days. Anxiety, general locomotor activity, willingness to explore, hippocampal and peri-postrhinal cortex dependent memory was investigated after eight weeks using Open field combined with Novel location/Novel object recognition tests. Toxin exposed animals did not perform worse than controls, and MC-LR exposed animals performed somewhat better during the first Barnes maze re-test session. MC-LR exposed mice rapidly lost up to ~5% body weight, but regained weight from day eight.”

Reference:

Oddvar Myhre, Dag Marcus Eide, Synne Kleiven, Hans Christian Utkilen & Tim Hofer (2018), Scientific Reports 8, Article number: 2308. doi:10.1038/s41598-018-20327-y

 

Metabolic solutions to the biosynthesis of BMAA and 2,4-DAB in cyanobacteria

The pathways of BMAA (and the related 2,4-DAB) biosynthesis in cyanobacteria are not known. A recent review by Peter Nunn and Geoffrey Codd considers possible metabolic routes, by analogy with reactions used in other species, by which these amino acids might be biosynthesised by cyanobacteria, which are a widespread potential environmental source of these neurotoxins.

Reference: Nunn, P. B. and Codd, G. A. (2017). “Metabolic solutions to the biosynthesis of some diaminomonocarboxylic acids in nature: Formation in cyanobacteria of the neurotoxins 3-N-methyl-2,3-diaminopropanoic acid (BMAA) and 2,4-diaminobutanoic acid (2,4-DAB)”. Phytochemistry 144, 253-270.

https://doi.org/10.1016/j.phytochem.2017.09.015