5-HT7R/G12 signaling regulates neuronal morphology and function in an age-dependent manner.

The common neurotransmitter serotonin controls different aspects of early neuronal differentiation, although the underlying mechanisms are poorly understood. Here we report that activation of the serotonin 5-HT(7) receptor promotes synaptogenesis and enhances synaptic activity in hippocampal neurons at early postnatal stages. An analysis of Gα(12)-deficient mice reveals a critical role of G(12)-protein for 5-HT(7) receptor-mediated effects in neurons. In organotypic preparations from the hippocampus of juvenile mice, stimulation of 5-HT(7)R/G(12) signaling potentiates formation of dendritic spines, increases neuronal excitability, and modulates synaptic plasticity. In contrast, in older neuronal preparations, morphogenetic and synaptogenic effects of 5-HT(7)/G(12) signaling are abolished. Moreover, inhibition of 5-HT(7) receptor had no effect on synaptic plasticity in hippocampus of adult animals. Expression analysis reveals that the production of 5-HT(7) and Gα(12)-proteins in the hippocampus undergoes strong regulation with a pronounced transient increase during early postnatal stages. Thus, regulated expression of 5-HT(7) receptor and Gα(12)-protein may represent a molecular mechanism by which serotonin specifically modulates formation of initial neuronal networks during early postnatal development.

Utilization of iPSC-derived human neurons for high-throughput drug-induced peripheral neuropathy screening.

As the number of cancer survivors continues to grow, awareness of long-term toxicities and impact on quality of life after chemotherapy treatment in cancer survivors has intensified. Chemotherapy-induced peripheral neuropathy (CIPN) is one of the most common side effects of modern chemotherapy. Animal models are used to study peripheral neuropathy and predict human risk; however, such models are labor-intensive and limited translatability between species has become a major challenge. Moreover, the mechanisms underlying CIPN have not been precisely determined and few human neuronal models to study CIPN exist. Here, we have developed a high-throughput drug-induced neurotoxicity screening model using human iPSC-derived peripheral-like neurons to study the effect of chemotherapy agents on neuronal health and morphology using high content imaging measurements (neurite length and neuronal cell viability). We utilized this model to test various classes of chemotherapeutic agents with known clinical liability to cause peripheral neuropathy such as platinum agents, taxanes, vinca alkaloids, proteasome inhibitors, and anti-angiogenic compounds. The model was sensitive to compounds that cause interference in microtubule dynamics, especially the taxane, epothilone, and vinca alkaloids. Conversely, the model was not sensitive to platinum and anti-angiogenic chemotherapeutics; compounds that are not reported to act directly on neuronal processes. In summary, we believe this model has utility for high-throughput screening and prediction of human risk for CIPN for novel chemotherapeutics.

Signaling mechanisms of metabotropic glutamate receptor 5 subtype and its endogenous role in a locomotor network.

Metabotropic glutamate receptors (mGluRs) act as modulators in the CNS of vertebrates, but their role in motor pattern generation in particular is primarily unknown. The intracellular signaling mechanisms of the group I mGluRs (mGluR1 and mGluR5), and their endogenous role in regulating locomotor pattern generation have been investigated in the spinal cord of the lamprey. Application of the group I mGluR agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) produced oscillations of the intracellular Ca2+ concentration ([Ca2+]i) in neurons. The oscillations were blocked by the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP) but not by the mGluR1 antagonist 7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxylate ethyl ester. These [Ca2+]i oscillations were abolished by a phospholipase C blocker and after depletion of internal Ca2+ stores by thapsigargin but did not involve protein kinase C activation. Furthermore, they were dependent on Ca2+ influx, because no [Ca2+]i oscillations were produced by DHPG in a Ca2+-free solution or after blockade of L-type Ca2+ channels. The mGluR5 is activated by an endogenous release of glutamate during locomotion, and a receptor blockade by MPEP caused an increase in the burst frequency. Thus, our results show that mGluR5 induces [Ca2+]i oscillations and regulates the activity of locomotor networks through endogenous activation.

Cardiac safety pharmacology: from human ether-a-gogo related gene channel block towards induced pluripotent stem cell based disease models.

INTRODUCTION: The field of cardiac safety pharmacology has been experiencing exciting changes over the recent years. Drug induced arrhythmia of the torsade des pointes types has been the reason for the denial of approval of novel drug candidates. The aim of cardiac safety pharmacology is to detect undesirable pharmacodynamic drug effects within and above the therapeutic range. A special focus is on the identification of potential arrhythmogenic effects within the drug discovery chain. AREAS COVERED: Here, the authors discuss the relevance of induced pluripotent stem (iPS) cell derived cardiomyocytes for safety pharmacology. The technology of obtaining functional cardiomyocytes from somatic cells of healthy donors and patients with inherited diseases is the basis for diverse disease models in multi-level safety pharmacology screening. The reader will gain an overview of stem cell based technologies in cardiac safety pharmacology in cardiac and disease modeling by iPS cell derived cardiomyocytes from patients with an inherited cardiac syndrome. EXPERT OPINION: iPS cell derived cardiomyocytes - especially from patients with increased risk of cardiac arrhythmia - are on the verge of offering new options for drug testing. More reliable assays can be expected to predict the arrhythmogenic risk of drug candidates in humans. However, this technology is still new and extensive validation studies are due.

Tetrahydrochromenoimidazoles as potassium-competitive acid blockers (P-CABs): structure-activity relationship of their antisecretory properties and their affinity toward the hERG channel.

Potassium-competitive acid blockers (P-CABs) constitute a new therapeutic option for the treatment of acid-related diseases that are widespread and constitute a significant economical burden. Enantiomerically pure tetrahydrochromenoimidazoles were prepared using the readily available candidate 4 (BYK 405879) as starting material or the Noyori asymmetric reduction of ketones as key reaction. A comprehensive SAR regarding the influence of the 5-carboxamide and the 8-aryl residue on in vitro activity, acid-suppression in the Ghosh Schild rat, and affinity toward the hERG channel was established. In addition, efficacy and duration of the antisecretory action was examined for the most promising target compounds by 24 h pH-metry in the fistula dog and a significantly different SAR was observed as compared to the Ghosh Schild rat. Several tetrahydrochromenoimidazoles were identified that possessed a comparable profile as the candidate 4.

Characterization of Na+-activated K+ currents in larval lamprey spinal cord neurons.

Potassium channels play an important role in controlling neuronal firing and synaptic interactions. Na(+)-activated K(+) (K(Na)) channels have been shown to exist in neurons in different regions of the CNS, but their physiological function has been difficult to assess. In this study, we have examined if neurons in the spinal cord possess K(Na) currents. We used whole cell recordings from isolated spinal cord neurons in lamprey. These neurons display two different K(Na) currents. The first was transient and activated by the Na(+) influx during the action potentials, and it was abolished when Na(+) channels were blocked by tetrodotoxin. The second K(Na) current was sustained and persisted in tetrodotoxin. Both K(Na) currents were abolished when Na(+) was substituted with choline or N-methyl-D-glucamine, indicating that they are indeed dependent on Na(+) influx into neurons. When Na(+) was substituted with Li(+), the amplitude of the inward current was unchanged, whereas the transient K(Na) current was reduced but not abolished. This suggests that the transient K(Na) current is partially activated by Li(+). These two K(Na) currents have different roles in controlling the action potential waveform. The transient K(Na) appears to act as a negative feedback mechanism sensing the Na(+) influx underlying the action potential and may thus be critical for setting the amplitude and duration of the action potential. The sustained K(Na) current has a slow kinetic of activation and may underlie the slow Ca(2+)-independent afterhyperpolarization mediated by repetitive firing in lamprey spinal cord neurons.

Metabotropic glutamate receptors provide intrinsic modulation of the lamprey locomotor network.

Spinal networks generate the motor pattern underlying locomotion. These are subject to modulatory systems that influence their operation and thereby result in a flexible network organization. In this review, we have summarized the mechanisms by which the different metabotropic glutamate receptor subtypes fine-tune the cellular and synaptic properties and thus underlie intrinsic modulation of the activity of the locomotor network in the lamprey.

mGluR1, but not mGluR5, mediates depolarization of spinal cord neurons by blocking a leak current.

The modulation of neuronal excitability by group I metabotropic glutamate receptors (mGluRs) was studied in isolated lamprey spinal cord. At resting potential, application of the group I mGluR agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) slightly depolarized the cells. However, at depolarized membrane potentials, this agonist induced repetitive firing. When Na+ channels were blocked by TTX, DHPG induced a slight depolarization at rest that increased in amplitude as the neurons were held at more depolarized membrane potentials. In voltage-clamp conditions, DHPG application induced an inward current associated with a decrease in membrane conductance when cells were held at -40 mV. At resting membrane potential, no significant change in the current was induced by DHPG, although a decrease in membrane conductance was seen. The conductance blocked by DHPG corresponded to a leak current, since DHPG had no effect on the voltage-gated current elicited by a voltage step from -60 to -40 mV, when leak currents were subtracted. The leak current blocked by DHPG is mediated by fluxes of both K+ and Na+. The subtype of group I mGluR mediating the block of the leak current was characterized using specific antagonists for mGluR1 and mGluR5. The inhibition of the leak current was blocked by the mGluR1 antagonist LY 367385 but not by the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP). The DHPG-induced blockage of the leak current required phospholipase C (PLC)-activation and release of Ca2+ from internal stores as the effect of DHPG was suppressed by the PLC-blocker U-73122 and after depletion of intracellular Ca2+ pools by thapsigargin. Our results thus show that mGluR1 activation depolarizes spinal neurons by inhibiting a leak current. This will boost membrane depolarization and result in an increase in the excitability of spinal cord neurons, which could contribute to the modulation of the activity of the spinal locomotor network.