No abstract

Depolarization drives neuronal plasticity. However, whether depolarization drives sensitization of peripheral nociceptive neurons remains elusive. By high-content screening (HCS) microscopy, we revealed that depolarization of cultured sensory neurons rapidly activates protein kinase A type II (PKA-II) in nociceptors by calcium influx through CaV1.2 channels. This effect was modulated by calpains but insensitive to inhibitors of cAMP formation, including opioids. In turn, PKA-II phosphorylated Ser1928 in the distal C terminus of CaV1.2, thereby increasing channel gating, whereas dephosphorylation of Ser1928 involved the phosphatase calcineurin. Patch-clamp and behavioral experiments confirmed that depolarization leads to calcium- and PKA-dependent sensitization of calcium currents ex vivo and local peripheral hyperalgesia in the skin in vivo. Our data suggest a local activity-driven feed-forward mechanism that selectively translates strong depolarization into further activity and thereby facilitates hypersensitivity of nociceptor terminals by a mechanism inaccessible to opioids.

BackgroundDiagnostic yield in patients with autism spectrum disorder (ASD) has improved over the last years, thanks to the introduction of whole genome arrays and next generation sequencing, but etiology is still unknown for the majority of cases. Among distinct cellular pathways, evidence implicating dysregulation of cellular calcium homeostasis in ASD pathogenesis has been accumulating, and speci c mutations in voltagegated calcium channels found in patients with autism were shown to be functionally relevant. MethodsWhole exome sequencing and Sanger sequencing were performed to identify and con rm variants in a girl with ASD, global developmental delay and precocious puberty, born of rst-degree cousins. Sitedirected mutagenesis was used to generate a human Ca V β 2d calcium channel subunit carrying a CACNB2 mutation. Whole-cell patch-clamp recordings were performed to reveal functional effects of mutant Ca V β 2d on Ba 2+ -currents mediated by L-type (Ca V 1.2) calcium channels in transiently transfected HEK-293 cells. ResultsIn an ASD patient, we identi ed a rare homozygous variant (p.Arg70Cys) in the CACNB2 gene coding for the auxiliary Ca V β 2 subunit of voltage-gated calcium channels. In a recombinant system, the Ca V β 2 variant, which was not previously associated to ASD, was found to alter Ca V 1.2 calcium channel function by signi cantly affecting activation and inactivation of whole-cell Ba 2+ -currents. LimitationsAlthough the evidence of CACNB2 involvement in ASD is slowly accumulating, the number of reported patients is very limited. Deep clinical phenotyping and functional studies in larger sets of subjects will be instrumental to fully understand the penetrance and outcome of CACNB2 variants. ConclusionsThe p.Arg70Cys variant in CACNB2 shows functional consequences similar to other ASD-associated Ca V β 2 mutations. These results support the idea of CACNB2 variations contributing to the development of ASD and hint to a rare form of Mendelian recessive autism with possible speci c comorbidities.

Voltage-gated calcium channel (VGCC) subunits have been genetically associated with autism spectrum disorders (ASD). The properties of the pore-forming VGCC subunit are modulated by auxiliary β-subunits, which exist in four isoforms (CaVβ1-4). Our previous findings suggested that activation of L-type VGCCs is a common feature of CaVβ2 subunit mutations found in ASD patients. In the current study, we functionally characterized a novel CaVβ1b variant (p.R296C) identified in an ASD patient. We used whole-cell and single-channel patch clamp to study the effect of CaVβ1b_R296C on the function of L- and N-type VGCCs. Furthermore, we used co-immunoprecipitation followed by Western blot to evaluate the interaction of the CaVβ1b-subunits with the RGK-protein Gem. Our data obtained at both, whole-cell and single-channel levels, show that compared to a wild-type CaVβ1b, the CaVβ1b_R296C variant inhibits L- and N-type VGCCs. Interaction with and modulation by the RGK-protein Gem seems to be intact. Our findings indicate functional effects of the CaVβ1b_R296C variant differing from that attributed to CaVβ2 variants found in ASD patients. Further studies have to detail the effects on different VGCC subtypes and on VGCC expression.

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Zusammenfassung Hintergrund Herz-Kreislauf-Erkrankungen sind nach wie vor die häufigste Todesursache weltweit. Neben einem erhöhten Blutdruck ist ein weiterer modifizierbarer Risikofaktor ein erhöhtes Low-density-Lipoprotein-Cholesterin. Obwohl beides gut medikamentös kontrollierbar ist, bleibt die Kontrolle bisher mangelhaft. Eine wesentliche Ursache ist eine unzureichende Adhärenz zur Medikation. Eine Lösung hierfür ist das Konzept der „Polypill“, also die Kombination mehrerer Wirkstoffe in einer einzelnen Tablette. Hierdurch wird nicht nur die Therapieadhärenz verbessert, sondern auch eine Verringerung kardiovaskulärer Ereignisse und eine Verbesserung der Prognose der Patienten erreicht. Ziel der Übersicht Diese Übersichtsarbeit fasst die aktuellen Evidenzen aus randomisierten klinischen Studien in der Primär- und Sekundärprävention zusammen. Ein wesentlicher Fokus liegt auf der aktuell publizierten SECURE-Studie, die die Wirksamkeit der „Polypill“ in der Sekundärprävention untersucht. Datenlage Viele Studien zur „Polypill“ beschäftigen sich mit der Kontrolle der Risikofaktoren und der Verbesserung der Therapieadhärenz, ohne jedoch einen prognostischen Vorteil zu adressieren. Neuere Studien wie HOPE‑3, PolyIran und TIPS‑3 konnten in der Primärprävention einen prognostischen Vorteil aufzeigen. In der Sekundärprävention war dies bis jetzt noch nicht geschehen. Diese Lücke wurde nun durch die SECURE-Studie geschlossen. Hier wurde bei Patienten nach Infarkt nicht nur eine signifikante Reduktion schwerwiegender kardiovaskulärer Ereignisse, sondern auch eine Reduktion kardiovaskulärer Todesfälle durch die „Polypill“ nachgewiesen. Schlussfolgerung Das Konzept der „Polypill“ hat sich von einer Komfortmaßnahme – einer Erleichterung der Medikamenteneinnahme für die Patienten – weiterentwickelt hin zu einem innovativen Therapiekonzept mit nachgewiesenem prognostischem Vorteil in Form einer Reduktion schwerwiegender Ereignisse und Todesfälle. Es ist an der Zeit, das Konzept der „Polypill“ breit einzusetzen, um die Bürde der Herz-Kreislauf-Erkrankungen weltweit zur verringern.

BackgroundDiagnostic yield in patients with autism spectrum disorder (ASD) has improved over the last years, thanks to the introduction of whole genome arrays and next generation sequencing, but etiology is still unknown for the majority of cases. Among distinct cellular pathways, evidence implicating dysregulation of cellular calcium homeostasis in ASD pathogenesis has been accumulating, and specific mutations in voltage-gated calcium channels found in patients with autism were shown to be functionally relevant.MethodsWhole exome sequencing and Sanger sequencing were performed to identify and confirm variants in a girl with ASD, global developmental delay and precocious puberty, born of first-degree cousins. Site-directed mutagenesis was used to generate a human CaVβ2d calcium channel subunit carrying a CACNB2 mutation. Whole-cell patch-clamp recordings were performed to reveal functional effects of mutant CaVβ2d on Ba2+-currents mediated by L-type (CaV1.2) calcium channels in transiently transfected HEK-293 cells.ResultsIn an ASD patient, we identified a rare homozygous variant (p.Arg70Cys) in the CACNB2 gene coding for the auxiliary CaVβ2subunit of voltage-gated calcium channels. In a recombinant system, the CaVβ2 variant, which was not previously associated to ASD, was found to alter CaV1.2 calcium channel function by significantly affecting activation and inactivation of whole-cell Ba2+-currents.LimitationsAlthough the evidence of CACNB2 involvement in ASD is slowly accumulating, the number of reported patients is very limited. Deep clinical phenotyping and functional studies in larger sets of subjects will be instrumental to fully understand the penetrance and outcome of CACNB2 variants.ConclusionsThe p.Arg70Cys variant in CACNB2 shows functional consequences similar to other ASD-associated CaVβ2 mutations. These results support the idea of CACNB2 variations contributing to the development of ASD and hint to a rare form of Mendelian recessive autism with possible specific comorbidities.