Ca²⁺-Induced Ca²⁺Release inAplysiaBag Cell Neurons Requires Interaction Between Mitochondrial and Endoplasmic Reticulum Stores

Abstract

Intracellular Ca2+ is influenced by both Ca2+ influx and release. We examined intracellular Ca2+ following action potential firing in the bag cell neurons of Aplysia californica. Following brief synaptic input, these neuroendocrine cells undergo an afterdischarge, resulting in elevated Ca2+ and the secretion of neuropeptides to initiate reproduction. Cultured bag cell neurons were injected with the Ca2+ indicator, fura-PE3, and subjected to simultaneous imaging and electrophysiology. Delivery of a 5-Hz, 1-min train of action potentials (mimicking the fast phase of the afterdischarge) produced a Ca2+ rise that markedly outlasted the initial influx, consistent with Ca2+-induced Ca2+ release (CICR). This response was attenuated by about half with ryanodine or depletion of the endoplasmic reticulum (ER) by cyclopiazonic acid. However, depletion of the mitochondria, with carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone, essentially eliminated CICR. Dual depletion of the ER and mitochondria did not reduce CICR further than depletion of the mitochondria alone. Moreover, tetraphenylphosphonium, a blocker of mitochondrial Ca2+ release, largely prevented CICR. The Ca2+ elevation during and subsequent to a stimulus mimicking the full afterdischarge was prominent and enhanced by protein kinase C activation. Traditionally, the ER is seen as the primary Ca2+ source for CICR. However, bag cell neuron CICR represents a departure from this view in that it relies on store interaction, where Ca2+ released from the mitochondria may in turn liberate Ca2+ from the ER. This unique form of CICR may be used by both bag cell neurons, and other neurons, to initiate secretion, activate channels, or induce gene expression.

Alternative oxidase in animals: unique characteristics and taxonomic distribution

“…The starlet sea anemone, N. vectensis, has rapidly emerged as a model system for the investigation of gene families and developmental patterning in animals (Matus et al, 2008). The sea hare, A. californica, is used to study the central nervous system and memory formation (Geiger and Magoski, 2008;Hawkins et al, 2006). Branchiostoma floridae (amphioxus) has been used in gene evolution studies, especially those examining homeobox genes involved in embryonic development (Takatori et al, 2008).…”

Ca2+removal by the plasma membrane Ca2+-ATPase influences the contribution of mitochondria to activity-dependent Ca2+dynamics inAplysianeuroendocrine cells

“…(CICR) from intracellular organelles (Berridge 2002). Although CICR has classically been described as arising from the endoplasmic reticulum (ER), an analogous mechanism can also be mediated by the mitochondria and exists in a variety of vertebrate and invertebrate neuronal types (Colegrove et al 2000a;Friel and Tsien 1994;Geiger and Magoski 2008;Groten et al 2013;Lee et al 2007;Tang and Zucker 1997;Werth and Thayer 1994). This process is initiated when Ca 2ϩ derived from voltage-gated Ca 2ϩ channels diffuses into the mitochondria through a Ca 2ϩ -selective ion channel on the inner mitochondrial membrane-the mitochondrial Ca 2ϩ uniporter (MCU) (Baughman et al 2011;Kirichok et al 2004).…”

“…During this time, egg-laying hormone is released to initiate the highpriority fixed action pattern of egg laying behavior (Arch 1972;Loechner et al 1990; Pinsker and Dudek 1977;Wayne et al 1998). Prior work has established that the mitochondria contribute prominently to both Ca 2ϩ removal and CICR after voltage-gated Ca 2ϩ influx in cultured bag cell neurons (Geiger and Magoski 2008;Groten et al 2013). Consequently, the mitochondria are implicated in sustaining peptide release and membrane excitability during the afterdischarge (Fisher et al 1994;Hickey et al 2010Hickey et al , 2013Michel and Wayne 2002;Wayne et al 1998).…”

Electrical coupling betweenAplysiabag cell neurons: characterization and role in synchronous firing

“…The maximal final concentration of DMSO or ethanol ranged from 0.05 to 0.5% (vol/vol), which in control experiments had no effect on holding current, membrane conductance, or junctional current. Prior work from our laboratory has found that these vehicles do not alter bag cell neuron macroscopic or single-channel currents, membrane potential, membrane capacitance, or intracellular Ca 2ϩ (Magoski et al 2000;Magoski and Kaczmarek 2005;Kachoei et al 2006;Lupinsky and Magoski 2006;Hung and Magoski 2007;Gardam et al 2008;Geiger and Magoski 2008;Tam et al 2009Tam et al , 2011Hickey et al 2010Hickey et al , 2013. We also attempted to employ carbenoxolone (C4790; Sigma-Aldrich) but were unable to dissolve it into nASW from either a DMSO-or ethanol-based stock.…”