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Background: Delta-opioid receptor (DOR)-mediated modulation of hippocampal neural networks is involved in emotions, cognition, and in pathophysiology and treatment of mood disorders. In this study, we examined the effects of DOR agonist (SNC80) and antagonist (naltrindole) on the excitability of individual hippocampal neurons., Methods: Primary neuronal cultures were prepared from hippocampi of newborn rats and cultivated in vitro for 8-14 days (DIV8-14). The effects of SNC80 naltrindole on evoked and spontaneous action potentials (APs) were measured at DIV8-9 and DIV13-14, respectively., Results: SNC80 (100 µM) potentiated spontaneous AP firing and stimulated sodium current; naltrindole had opposite effects. The stimulatory effect of 100 µM of SNC80 was revoked by pre-administration of 1 µM of naltrindole. SNC80 and naltrindole induced similar inhibitory effects on the evoked AP firing and on the calcium current. Further, SNC80 inhibited both peak and sustained potassium currents. Naltrindole had no effect on potassium currents., Conclusion: We suggest that the effects of naltrindole and high concentration of SNC80 on the sodium currents are mediated via DORs and underlying the changes in spontaneous activity. The inhibitory effects of SNC80 on calcium and potassium currents might also be DOR-dependent; these currents might mediate SNC80 effect on the evoked AP firing. The inhibitory effects of naltrindole on calcium and of low doses of SNC80 on sodium currents might be however DOR independent. The behavioral effects of SNC80 and naltrindole, observed in previous studies, might be mediated, at least in part, via the modulatory effect of these ligands on the excitability of hippocampal neurons.

Voltage-gated Ca 2+ channels are typically integrated in a complex network of protein-protein-interactions, also referred to as Ca 2+ channel nanodomains. Amongst the neuronal Ca V 2 channel family, Ca V 2.2 is of particular importance due to its general role for signal transmission from the periphery to the central nervous system, but also due to its significance for pain perception. Thus, Ca V 2.2 is an ideal target candidate to search for pharmacological inhibitors but also for novel modulatory interactors. In this review we summarize the last years findings of our intense screenings and characterization of the six Ca V 2.2 interaction partners, tetraspanin-13 (TSPAN-13), reticulon 1 (RTN1), member 1 of solute carrier family 38 (SLC38), prostaglandin D2 synthase (PTGDS), transmembrane protein 223 (TMEM223), and transmembrane BAX inhibitor motif 3 (Grina/TMBIM3) containing protein. Each protein shows a unique way of channel modulation as shown by extensive electrophysiological studies. Amongst the newly identified interactors, Grina/TMBIM3 is most striking due to its modulatory effect which is rather comparable to G-protein regulation.

Traumatic injury of the central nervous system is accompanied by various functional and morphological changes. Animal models of traumatic brain injury are commonly used to investigate changes in behaviour, morphology, in the expression of various proteins around the site of the injury, or the expression of diagnostically important biomarkers. Excitability of a single neuron at, or close to, the site of injury was rarely investigated. Several in vitro models were developed which allow such investigation. In the present work, we employed a fibrotic scar model according to Kimura-Kuroda and coauthors to analyse altered excitability of rat hippocampal neurons under the conditions mimicking traumatic brain injury. Hippocampal neurons from newborn rats were cultured either on a fibrotic scar model or in the presence of TGF-β1, a cytokine secreted at a brain injury site that may have both neuroprotective and neurodegenerative function. Fibrotic scar facilitated ability of neonatal hippocampal neurons to fire action potential series by increasing the density of voltage activated sodium and potassium currents. Chondroitin sulphate proteoglycans played substantial role in these effects, as proven by their full reversion after administration of Chondroitinase ABC. In contrast, TGF-β1 did not contribute to them. An application of TGF-β1 itself attenuated generation of action potentials, inhibited sodium current and potentiated potassium currents. Main alteration of electrophysiological parameters of neonatal hippocampal neurons caused by a fibrotic scar model is enhanced excitability. TGF-β1 may have predominantly neuroprotective role in injured rat hippocampus., (© 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Acute injury to central nervous system (CNS) triggers neurodegenerative processes that can result in serious damage or complete loss of function. After injury, production of transforming growth factor β1 (TGFβ1) increases and initiates creation of a fibrotic scar that prevents normal growth, plasticity, and recovery of damaged neurons. Administration of TGFβ1 antagonists can prevent its pathological effects. To define consequences of increased TGFβ1 release on calcium signaling, neuronal plasticity, excitability, and mitochondrial dynamics in CNS neurons we directly exposed a rat primary culture of cerebellar granule neurons to TGFβ1. We focused on changes in expression of intracellular calcium transporters, especially inositol-1,4,5-trisphosphate receptor (IP3R) type 1, mitochondrial dynamics, and membrane excitability. TGFβ1 significantly decreased the gene and protein expression of inositol-1,4,5-trisphosphate receptor type 1 and the gene expression of additional intracellular Ca transporters such as IP3R2, ryanodine receptor type 1 (RyR1), RyR2, and SERCA2. Altered calcium signaling suppressed neurite outgrowth and significantly decreased the length of the mitochondria and the frequency of mitochondrial fusion. The resting membrane potential of cerebellar granule neurons was hyperpolarized and slow after depolarization of single action potential was suppressed. LY364947, a blocker of TGFβ1 receptor I, prevented these effects, and IP3 receptor blocker 2-aminoethoxydiphenyl borate (2APB) mimicked them. After CNS injury TGFβ1 downregulates intracellular Ca levels and alters Ca signaling within injured neurons. We suggest that in our model TGFβ1 may trigger both neurodegenerative and neuroprotective events through IP3-induced Ca signaling.

L-type calcium channels (LTCC) are important functional elements of hippocampal neurons contributing to processes like memory formation and gene expression. Mice lacking the Ca(V)1.2 channel in hippocampal pyramidal cells exhibited defects in spatial memory (Moosmang et al. 2005) and lowered frequency of repetitive action potential (AP) firing (Lacinova et al. 2008). We tested the contribution of LTCC to AP firing of cultured rat neonatal hippocampal neurons using the dihydropyridine channel blocker nimodipine. Ionic currents and APs were recorded in the whole cell patch clamp configuration. A prolonged depolarizing current pulse activated the firing of a series of APs. The presence of 10 μM nimodipine blocked all but the first AP in series. This concentration, which is potent enough to completely block LTCC, inhibited about 35-50% of the total calcium current. In addition, nimodipine blocked about 50% of both calcium-dependent and voltage-dependent potassium currents whereas the sodium current was not affected. We suggest that nimodipine suppressed the firing of APs in cultured neonatal rat hippocampal neurons due to inhibition of both calcium and potassium currents.

The auxiliary calcium channel alpha2delta subunit comprises a family of three genes, alpha2delta-1 to 3, which are expressed in a tissue-specific manner. alpha2delta-2 mRNA is found in the heart, skeletal muscle, brain, kidney, liver and pancreas. We report here for the first time the identification and functional characterization of alpha2delta-2 splice variants and their mRNA distribution in the mouse brain. The splice variants differ in the alpha2 and delta protein by eight and three amino acid residues, respectively, and are differentially expressed in cardiac tissue and human medullary thyroid carcinoma (hMTC) cells. In situ hybridization of mouse brain sections revealed the highest expression of alpha2delta-2 mRNA in the Purkinje cell layer of the cerebellum, habenulae and septal nuclei, and a lower expression in the cerebral cortex, olfactory bulb, thalamic and hypothalamic nuclei, as well as the inferior and superior colliculus. As the in situ data did not suggest a specific colocalization with any alpha1 subunit, coexpression studies of alpha2delta-2 were carried out either with the high-voltage-gated calcium channels, alpha1C, alpha1E or alpha1A, or with the low-voltage-gated calcium channel, alpha1G. Coexpression of alpha2delta-2 increased the current density, shifted the voltage dependence of channel activation and inactivation of alpha1C, alpha1E and alpha1A subunits in a hyperpolarizing direction, and accelerated the decay and shifted the steady-state inactivation of the alpha1G current.

Abstract The auxiliary calcium channel α2δ subunit comprises a family of three genes, α2δ-1 to 3, which are expressed in a tissue-specific manner. α2δ-2 mRNA is found in the heart, skeletal muscle, brain, kidney, liver and pancreas. We report here for the first time the identification and functional characterization of α2δ-2 splice variants and their mRNA distribution in the mouse brain. The splice variants differ in the α2 and δ protein by eight and three amino acid residues, respectively, and are differentially expressed in cardiac tissue and human medullary thyroid carcinoma (hMTC) cells. In situ hybridization of mouse brain sections revealed the highest expression of α2δ-2 mRNA in the Purkinje cell layer of the cerebellum, habenulae and septal nuclei, and a lower expression in the cerebral cortex, olfactory bulb, thalamic and hypothalamic nuclei, as well as the inferior and superior colliculus. As the in situ data did not suggest a specific colocalization with any α1 subunit, coexpression studies of α2δ-2 were carried out either with the high-voltage-gated calcium channels, α1C, α1E or α1A, or with the low-voltage-gated calcium channel, α1G. Coexpression of α2δ-2 increased the current density, shifted the voltage dependence of channel activation and inactivation of α1C, α1E and α1A subunits in a hyperpolarizing direction, and accelerated the decay and shifted the steady-state inactivation of the α1G current. [ABSTRACT FROM AUTHOR]