A number of different cell lines that exhibit a partial neuronal phenotype have been identified, but in many cases the full extent of their neuronal differentiation has not been directly addressed by functional studies. We have used electrophysiology and immunofluorescence to examine the formation of synapses and the development of neuronal polarity by murine embryonic stem (ES) cells and the mouse P19 embryonic carcinoma cell line. Within 2-3 weeks after induction by retinoic acid, subsets of P19 and ES cells formed excitatory synapses, mediated by glutamate receptors, or inhibitory synapses, mediated by receptors for GABA or glycine. In ES-cell cultures, both NMDA and non-NMDA receptors contributed to the excitatory postsynaptic response. Staining with antibodies to growth-associated protein-43 and microtubule-associated protein-2 revealed segregation of immunoreactivity into separate axonal and somato-dendritic compartments, respectively. Consistent with our physiological evidence for synapse formation, intense punctate staining was observed with antibodies to the synaptic vesicle proteins synapsin, SV2, and synaptophysin. These results demonstrate the in vitro acquisition by pluri-potent cell lines of neuronal polarity and functional synaptic transmission that is characteristic of CNS neurons.

Members of the transforming growth factor‐β superfamily, including bone morphogenetic protein 4 (BMP‐4), have been implicated as regulators of neuronal and glial differentiation. To test for a possible role of BMP‐4 in early mammalian neural specification, we examined its effect on neurogenesis in aggregate cultures of mouse embryonic stem (ES) cells. Compared to control aggregates, in which up to 20% of the cells acquired immunoreactivity for the neuron‐specific antibody TuJ1, aggregates maintained for 8 days in serum‐free medium containing BMP‐4 generated 5‐ to 10‐fold fewer neurons. The action of BMP‐4 was dose dependent and restricted to the fifth through eighth day in suspension. In addition to the reduction in neurons, we observed that ES cell cultures exposed to BMP‐4 contained fewer cells that were immunoreactive for glial fibrillary acidic protein or the HNK‐1 neural antigen. Furthermore, under phase contrast, cultures prepared from BMP‐4–treated aggregates contained a significant proportion of nonneuronal cells with a characteristic flat, elongated morphology. These cells were immunoreactive for antibodies to the intermediate filament protein vimentin; they were rare or absent in control cultures. Treatment with BMP‐4 enhanced the expression of the early mesodermal genes brachyury and tbx6 but had relatively little effect on total cell number or cell death. Coapplication of the BMP‐4 antagonist noggin counteracted the effect of exogenous BMP‐4, but noggin alone had no effect on neuralization in either the absence or presence of retinoids. Collectively, our results suggest that BMP‐4 can overcome the neuralizing action of retinoic acid to enhance mesodermal differentiation of murine ES cells. © 1999 John Wiley & Sons, Inc. J Neurobiol 40: 271–287, 1999

No abstract

In seeking broad-spectrum anticonvulsants to treat epilepsy and other neurological disorders, we synthesized and tested a group of sulfamide derivatives (4a-k, 5), which led to the clinical development of 4a (JNJ-26990990). This compound exhibited excellent anticonvulsant activity in rodents against audiogenic, electrically induced, and chemically induced seizures, with very weak inhibition of human carbonic anhydrase-II (IC(50) = 110 microM). The pharmacological profile for 4a supports its potential in the treatment of multiple forms of epilepsy, including pharmacoresistant variants. Mechanistically, 4a inhibited voltage-gated Na(+) channels and N-type Ca(2+) channels but was not effective as a K(+) channel opener. The pharmacokinetics and metabolic properties of 4a are discussed.

Kutchukian, P. S. et al. (2017) Iterative focused screening with biological fingerprints identifies selective Asc-1 inhibitors distinct from traditional high throughput screening. ACS Chemical Biology, 12(2), pp. 519-527. (doi:10.1021/acschembio.6b00913) This is the author's final accepted version.There may be differences between this version and the published version. You are advised to consult the publisher's version if you wish to cite from it.http://eprints.gla.ac.uk/146530/