Stimulation of GABA-Induced Ca2+ Influx Enhances Maturation of Human Induced Pluripotent Stem Cell-Derived Neurons

Optimal use of patient-derived, induced pluripotent stem cells for modeling neuronal diseases is crucially dependent upon the proper physiological maturation of derived neurons. As a strategy to develop defined differentiation protocols that optimize electrophysiological function, we investigated the role of Ca²⁺ channel regulation by astrocyte conditioned medium in neuronal maturation, using whole-cell patch clamp and Ca²⁺ imaging. Standard control medium supported basic differentiation of induced pluripotent stem cell-derived neurons, as assayed by the ability to fire simple, single, induced action potentials. In contrast, treatment with astrocyte conditioned medium elicited complex and spontaneous neuronal activity, often with rhythmic and biphasic characteristics. Such augmented spontaneous activity correlated with astrocyte conditioned medium-evoked hyperpolarization and was dependent upon regulated function of L-, N- and R-type Ca²⁺ channels. The requirement for astrocyte conditioned medium could be substituted by simply supplementing control differentiation medium with high Ca²⁺ or γ-amino butyric acid (GABA). Importantly, even in the absence of GABA signalling, opening Ca²⁺ channels directly using Bay K8644 was able to hyperpolarise neurons and enhance excitability, producing fully functional neurons. These data provide mechanistic insight into how secreted astrocyte factors control differentiation and, importantly, suggest that pharmacological modulation of Ca²⁺ channel function leads to the development of a defined protocol for improved maturation of induced pluripotent stem cell-derived neurons.     

A Detailed Family History of Myocardial Infarction and Risk of Myocardial Infarction – A Nationwide Cohort Study

BackgroundFamily history of myocardial infarction (MI) is an independent risk factor for MI. Several genetic variants are associated with increased risk of MI and family history of MI in a first-degree relative doubles MI risk. However, although family history of MI is not a simple dichotomous risk factor, the impact of specific, detailed family histories has not received much attention, despite its high clinical relevance. We examined risk of MI by MIs in first- and second-degree relatives and by number and age of affected relatives.ConclusionA detailed family history, particularly number of affected first- and second-degree relatives, contributes meaningfully to risk assessment, especially in middle-aged persons. Future studies should test for potential improvement of risk algorithm prediction using detailed family histories.     

Review of the Eospilarctia yuennanica group (Lepidoptera, Erebidae, Arctiinae) from the Indo - Himalayan region, with description of two new species and one subspecies

Twonew Eospilarctia species and one new subspecies from China, Myanmar and Vietnam, respectively, are described. Superficially the new species Eospilarctia maciaisp. n., Eospilarctia naumannisp. n. and Eospilarctia yuennanica fansipanassp. n. resemble related congeners but they can be distinguished by the differences in wing pattern, genitalia and distribution provided. Eospilarctia yuennanica guangdonga Dubatolov, Kishida & Wang, 2008 is upgraded to species level. A checklist of the genus Eospilarctia and a key to the Eospilarctia yuennanica (Daniel, 1943) species-group, based on external characters and male genitalia, is presented.     

Novel aminoglycosides increase SMN levels in spinal muscular atrophy fibroblasts

Spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality. SMA is caused by the homozygous absence of survival motor neuron-1 (SMN1). SMN2, a nearly identical copy gene, is retained in all SMA patients and encodes an identical protein as SMN1; however, SMN1 and SMN2 differ by a silent C to T transition which results in the production of an alternatively spliced isoform (SMNΔ7), which encodes a defective protein, demonstrating that the absence of the short peptide encoded by SMN exon 7 is critical in SMA development. Previously, we have shown that for some functions heterologous sequences can compensate for the exon 7 peptide, suggesting that the SMN C-terminus functions non-specifically. Consistent with this hypothesis, we now identify novel aminoglycosides that can induce SMN protein levels in patient fibroblasts. This hypothesis was supported, in part, by a novel fluorescent SMN read-through assay. Interestingly, however, through the development of a SMN exon 7-specific antibody, results suggested that levels of normal full-length SMN might also be elevated by aminoglycoside treatment. These results demonstrate that the compounds that promote read-through may provide an alternative platform for the discovery of compounds that induce SMN protein levels.Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Potent oxindole based human beta3 adrenergic receptor agonists

The synthesis and biological evaluation of a series of oxindole beta(3) adrenergic receptor agonists is described. A modulation of rat atrial tachycardia was observed with substitution at the 3-position of the oxindole moiety.     

In vitro neuropharmacological evaluation of penitrem-induced tremorgenic syndromes: importance of the GABAergic system

The effects of the fungal neurotoxin penitrem A on the GABAergic and glutamatergic systems in rat brain were evaluated. Penitrem A inhibited binding of the GABA_A-receptor ligand [³H]TBOB to rat forebrain and cerebellar membrane preparations with IC₅₀ (half maximal inhibitory concentration) values of 11 and 9 μM, respectively. Furthermore, penitrem A caused a concentration-dependent increase of [³H]flunitrazepam and [³H]muscimol binding in rat forebrain, but not in cerebellar preparations. The stimulation of [³H]flunitrazepam binding by penitrem A was abolished by the addition of GABA. In cerebellar preparations, a different pharmacological profile was found, with penitrem A allosterically inhibiting [³H]TBOB binding by interacting with a bicuculline-sensitive site. Moreover, penitrem A inhibited the high affinity uptake of GABA and glutamate into cerebellar synaptosomes with IC₅₀ values of 20 and 47 μM, respectively. The toxin showed no effect on NMDA or AMPA glutamate receptor binding. In conclusion, our results suggest that penitrem A exerts region-specific effects in the brain, leading to positive modulation of GABA_A-receptor function in forebrain. Conversely, penitrem A may act as a bicuculline-like convulsant in cerebellum.     

Purification and characterization of bacteriophage P22 Xis protein

The temperate bacteriophages λ and P22 share similarities in their site-specific recombination reactions. Both require phage-encoded integrase (Int) proteins for integrative recombination and excisionase (Xis) proteins for excision. These proteins bind to core-type, arm-type, and Xis binding sites to facilitate the reaction. λ and P22 Xis proteins are both small proteins (λ Xis, 72 amino acids; P22 Xis, 116 amino acids) and have basic isoelectric points (for P22 Xis, 9.42; for λ Xis, 11.16). However, the P22 Xis and λ Xis primary sequences lack significant similarity at the amino acid level, and the linear organizations of the P22 phage attachment site DNA-binding sites have differences that could be important in quaternary intasome structure. We purified P22 Xis and studied the protein in vitro by means of electrophoretic mobility shift assays and footprinting, cross-linking, gel filtration stoichiometry, and DNA bending assays. We identified one protected site that is bent approximately 137 degrees when bound by P22 Xis. The protein binds cooperatively and at high protein concentrations protects secondary sites that may be important for function. Finally, we aligned the attP arms containing the major Xis binding sites from bacteriophages λ, P22, L5, HP1, and P2 and the conjugative transposon Tn916. The similarity in alignments among the sites suggests that Xis-containing bacteriophage arms may form similar structures.     

Description of two new species of Cossidae (Lepidoptera) from China

Two new Cossidaespecies from China's Zhejiang and Sichuan provinces are described. The new species Phragmataecia monikasp. n.and Patoptoformis rimsaitesp. n. superficially resemble related congeners but can be distinguished by differences in wing pattern, genitalia and distribution. Checklists of the genera Phragmataecia and Patoptoformis are presented.     

Principles for applying optogenetic tools derived from direct comparative analysis of microbial opsins

Diverse optogenetic tools have allowed versatile control over neural activity. Many depolarizing and hyperpolarizing tools have now been developed in multiple laboratories and tested across different preparations, presenting opportunities but also making it difficult to draw direct comparisons. This challenge has been compounded by the dependence of performance on parameters such as vector, promoter, expression time, illumination, cell type and many other variables. As a result, it has become increasingly complicated for end users to select the optimal reagents for their experimental needs. For a rapidly growing field, critical figures of merit should be formalized both to establish a framework for further development and so that end users can readily understand how these standardized parameters translate into performance. Here we systematically compared microbial opsins under matched experimental conditions to extract essential principles and identify key parameters for the conduct, design and interpretation of experiments involving optogenetic techniques.