Super-Resolution Microscopy of the Neuronal Calcium-Binding Proteins Calneuron-1 and Caldendrin

Hradsky, Johannes; Mikhaylova, Marina; Karpova, Anna; Kreutz, Michael R.; Zuschratter, Werner

doi:10.1007/978-1-62703-230-8_10

Cited by 25 publications

(15 citation statements)

References 42 publications

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“…Cells were isolated as described in Ref. 8 and plated at a confluence of 40,000 cells/ 0.32 cm 2 in 96-well plates for MAPK experiments and in 6-well plates for the other assays. Cells were maintained in Neurobasal medium supplemented with 2 mM L-glutamine, 100 units/ml penicillin/streptomycin, and 2% (v/v) B27 supplement (Gibco) in a 96-well plate for 12 days.…”

Section: Methodsmentioning

confidence: 99%

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons

Navarro

Aguinaga

Angelats

et al. 2016

Journal of Biological Chemistry

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The truncated non-signaling ghrelin receptor growth hormone secretagogue R1b (GHS-R1b) has been suggested to simply exert a dominant negative role in the trafficking and signaling of the full and functional ghrelin receptor GHS-R1a. Here we reveal a more complex modulatory role of GHS-R1b. Differential co-expression of GHS-R1a and GHS-R1b, both in HEK-293T cells and in striatal and hippocampal neurons in culture, demonstrates that GHS-R1b acts as a dual modulator of GHSR1a function: low relative GHS-R1b expression potentiates and high relative GHS-R1b expression inhibits GHS-R1a function by facilitating GHS-R1a trafficking to the plasma membrane and by exerting a negative allosteric effect on GHS-R1a signaling, respectively. We found a preferential G i/o coupling of the GHSR1a-GHS-R1b complex in HEK-293T cells and, unexpectedly, a preferential G s/olf coupling in both striatal and hippocampal neurons in culture. A dopamine D 1 receptor (D1R) antagonist blocked ghrelin-induced cAMP accumulation in striatal but not hippocampal neurons, indicating the involvement of D1R in the striatal GHS-R1a-G s/olf coupling. Experiments in HEK-293T cells demonstrated that D1R co-expression promotes a switch in GHS-R1a-G protein coupling from G i/o to G s/olf , but only upon co-expression of GHS-R1b. Furthermore, resonance energy transfer experiments showed that D1R interacts with GHS-R1a, but only in the presence of GHS-R1b. Therefore, GHS-R1b not only determines the efficacy of ghrelin-induced GHS-R1a-mediated signaling but also determines the ability of GHS-R1a to form oligomeric complexes with other receptors, promoting profound qualitative changes in ghrelin-induced signaling.Ghrelin is an orexigenic hormone, an internal signal for the animal to engage in food-directed behavior (1, 2). It is produced by stomach oxyntic cells, which provide plasma levels that fluctuate diurnally with a peak in the day and trough at night. Notably, oxyntic cells qualify as food-entrained oscillators, and ghrelin plasma levels increase during anticipated mealtimes and decrease after meals (1). These and other less well characterized central neuronal functions of ghrelin depend on its ability to cross the blood-brain barrier by still unclear mechanisms and reaching ghrelin receptors localized in specific brain areas, such as the hypothalamus, hippocampus, amygdala, mesencephalic dopaminergic regions, and striatum (2-4).Ghrelin acts on the class A G protein-coupled receptor known as growth hormone secretagogue (GHS) 5 receptor or GHS-R1a. Cells expressing GHS-R1a also express GHS-R1b, a truncated variant of GHS-R1a lacking transmembrane domains 6 and 7. Ghrelin does not bind and therefore does not signal through GHS-R1b (5), and the role of this truncated "receptor" on ghrelin-mediated signaling is just beginning to be understood. Evidence has been provided for the ability of GHS-R1a to homodimerize and heterodimerize with GHS-R1b, which might allow GHS-R1b to produce a dominant negative effect on GHSR1a signaling. Two different mechanisms have b...

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Section: Methodsmentioning

confidence: 99%

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons

Navarro

Aguinaga

Angelats

et al. 2016

Journal of Biological Chemistry

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“…For neuronal primary cultures, the striatum from mouse embryos (E19) was removed and the neurons were isolated as described by (Hradsky et al, 2013) and plated at a density of circa 120,000 cells/cm 2 .…”

Section: Hek-293t Cells and Primary Culturesmentioning

confidence: 99%

Angiotensin AT1 and AT2 receptor heteromer expression in the hemilesioned rat model of Parkinson’s disease that increases with levodopa-induced dyskinesia

Rivas‐Santisteban

Rodríguez‐Pérez

Muñoz

et al. 2020

Preprint

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Background/Aims: The renin-angiotensin system (RAS) is altered in Parkinson’s disease (PD), a disease due to substantia nigra neurodegeneration and whose dopamine-replacement therapy, using the precursor levodopa, leads to dyskinesias as the main side effect. Angiotensin AT1 and AT2 receptors, mainly known for their role in regulating water homeostasis and blood pressure and able to form heterodimers (AT1/2Hets), are present in the central nervous system. We assessed the functionality and expression of AT1/2Hets in Parkinson Disease (PD).Methods: Immunocytochemistry was used to analyze the colocalization between angiotensin receptors, bioluminescence resonance energy transfer was used to detect AT1/2Hets. Calcium and cAMP determination, MAPK activation and label-free assays were performed to characterize signaling in homologous and heterologous systems. Proximity ligation assays was used to quantify receptor expression in mouse primary cultures and in rat striatal sections.Results: We confirmed that AT1 and AT2 receptors form AT1/2Hets that are expressed in cells of the central nervous system. AT1/2Hets are novel functional units with particular signaling properties. Importantly, the coactivation of the two receptors in the heteromer reduces the signaling output of angiotensin. Remarkably, AT1/2Hets that are expressed in both striatal neurons and microglia show a cross-potentiation, i.e. candesartan, the antagonist of AT1 increases the effect of AT2 receptor agonists. In addition, the level of striatal expression increased in the unilateral 6-OH-dopamine lesioned rat PD model and was markedly higher in parkinsonian-like animals that did not become dyskinetic upon levodopa chronic administration if compared with expression in those that became dyskinetic.Conclusion: The results indicate that boosting the action of neuroprotective AT2 receptors using an AT1 receptor antagonist constitutes a promising therapeutic strategy in PD.

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“…The sections were immersed with increasing TDE concentrations in PBS buffer (10, 25, 50, 97%, 10 min each) and finally embedded in 97% TDE. STED imaging was performed by using a commercial two-channel STED microscope (Leica Microsystems) as described previously 61 . Briefly, the system consists of an inverted microscope (DMI 6000 CS with a Tandem Scanning System SP5) equipped with two pulsed excitation lasers of 532 and 635 nm (E70 ps pulse width, 80 MHz repetition rate; PicoQuant) and a Titanium saphire laser (Chameleon ultra II (E100 fs pulse width, 80 MHz repetition rate), Coherent) as depletion light source.…”

Section: Methodsmentioning

confidence: 99%

KCNQ5 K+ channels control hippocampal synaptic inhibition and fast network oscillations

et al. 2015

Self Cite

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KCNQ2 (Kv7.2) and KCNQ3 (Kv7.3) K þ channels dampen neuronal excitability and their functional impairment may lead to epilepsy. Less is known about KCNQ5 (Kv7.5), which also displays wide expression in the brain. Here we show an unexpected role of KCNQ5 in dampening synaptic inhibition and shaping network synchronization in the hippocampus. KCNQ5 localizes to the postsynaptic site of inhibitory synapses on pyramidal cells and in interneurons. Kcnq5 dn/dn mice lacking functional KCNQ5 channels display increased excitability of different classes of interneurons, enhanced phasic and tonic inhibition, and decreased electrical shunting of inhibitory postsynaptic currents. In vivo, loss of KCNQ5 function leads to reduced fast (gamma and ripple) hippocampal oscillations, altered gamma-rhythmic discharge of pyramidal cells and impaired spatial representations. Our work demonstrates that KCNQ5 controls excitability and function of hippocampal networks through modulation of synaptic inhibition.

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Super-Resolution Microscopy of the Neuronal Calcium-Binding Proteins Calneuron-1 and Caldendrin

Cited by 25 publications

References 42 publications

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons

Angiotensin AT1 and AT2 receptor heteromer expression in the hemilesioned rat model of Parkinson’s disease that increases with levodopa-induced dyskinesia

KCNQ5 K+ channels control hippocampal synaptic inhibition and fast network oscillations

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