Electrically Evoked Synaptosomal Amino Acid Transmitter Release in Human Brain in Alcohol Misuse

Abstract

Severe chronic alcohol misuse leads to neuropathological changes in human brain, with the greatest neuronal loss in the dorsolateral prefrontal cortex. In this region, GABA_A receptors are selectively upregulated, and show altered subunit expression profiles only in alcoholics without comorbid disease, whereas glutamate_NMDA subunit expression profiles are selectively downregulated only in alcoholics with comorbid cirrhosis of the liver. To determine whether these outcomes might be conditional on synaptic transmitter levels, evoked release was studied in well-characterized synaptosome suspensions preloaded with L-[³H]glutamate and [¹⁴C]GABA and stimulated electrically (±10 V contiguous square waves, 0.4 ms, 100 Hz, 1.5 min) with and without Ca²⁺. Stimulation elicited brief peaks of both radioisotopes that were larger in the presence of Ca²⁺ ions (p < 0.01). A repeat stimulus evoked a second, smaller (p < 0.01) peak. Ca²⁺-dependent L-[³H]glutamate release, but not [¹⁴C]GABA release, was higher overall in alcoholics than in controls (p < 0.05). With comorbid cirrhosis, L-[³H]glutamate release showed a graded response, whereas [¹⁴C]GABA release was lowest in noncirrhotic alcoholics. Release patterns did not differ between cortical regions, or between males and females. Neither age nor postmortem interval was a significant confounder. The released transmitters may differentially alter receptor profiles on postsynaptic cells.

Glutamate Signaling in Alcohol Abuse and Dependence

“…Such a finding suggests that the basal hyper-glutamatergic state produced by a history of alcohol exposure does not persist into protracted withdrawal, but this requires further investigation within other mesocorticolimbic structures, particularly as functional analysis of glutamate signaling in human postmortem brain tissue indicated higher stimulated glutamate release from dorsolateral prefrontal cortex in alcoholics than in nonalcoholic controls (Kuo & Dodd, 2011). Moreover, akin to findings from studies of alcohol-dependent rats (Zahr et al, 2009), proton magnetic resonance spectroscopy in human alcoholics revealed increases in glutamate in prefrontal cortex (Hermann et al, 2012) and ventral striatum (Bauer et al, 2013) after detoxification.…”

Exosomal tau with seeding activity is released from Alzheimer's disease synapses, and seeding potential is associated with amyloid beta

“…The styryl dye FM2-10 quantifies synaptosome depolarization in vitro Synaptosomes are resealed nerved terminals prepared from fresh brain tissue that contain mitochondria and maintain membrane potential, along with a full complement of receptor and transporter functions 38 . Functional endpoints such as LTP and glucose transport can also be measured in postmortem human synaptosomes, if prepared from tissue that is cryopreserved by mincing and slow freezing in isotonic sucrose [39][40][41][42][43] . In order to verify and quantify depolarization in postmortem AD synaptosomes, we used FM2-10, a styryl dye with a lipophilic tail that becomes fluorescent on insertion into membranes 44,45 .…”

Global Analysis of Protein Phosphorylation Regulation upon Stimulation of Exocytosis in the Nerve Terminal

“…However, most of the studies with electrical stimulation of synaptosomes have applied long lasting (20-30 min) pulses to investigate the metabolic responses of synaptosomes. There is very limited knowledge available regarding the neurotransmission release upon electrical stimulation (Kuo and Dodd, 2011).…”