Introduction Conditionally immortalised human neural progenitor cells (hNPCs) represent a robust way to obtain native neural cells to investigate physiological mechanisms in both health and disease. Expression of neurotransmitter receptors, signalling proteins and related proteins were assessed by q- and RT-PCR and complemented by Ca2+ imaging, electrophysiology and assessment of ERK signalling in response to neurotransmitter ligand application. Finally, differentiated neurons were assessed for their ability to form putative synapses and to respond to activity-dependent stimulation. Results Differentiation of CTX0E16 hNPCs predominately resulted in the generation of neurons expressing markers of cortical and glutamatergic (excitatory) fate, and with a typical polarized neuronal morphology. Gene expression analysis confirmed an upregulation in the expression of cortical, glutamatergic and signalling proteins following differentiation. CTX0E16 neurons exhibited Ca2+ and ERK1/2 responses following exogenous neurotransmitter application, and after 6 weeks displayed spontaneous Ca2+ transients and electrophysiological properties consistent with that of immature neurons. Differentiated CTX0E16 neurons also expressed a range MPO-IN-28 of Rabbit polyclonal to GST pre- and post-synaptic proteins that co-localized along distal dendrites, and moreover, displayed structural plasticity in response to modulation of neuronal activity. Conclusions Taken together, these findings demonstrate that this CTX0E16 hNPC line is certainly a robust way to obtain cortical neurons, which screen functional properties in keeping with a glutamatergic phenotype. CTX0E16 neurons may be used to research cortical cell function Hence, and furthermore, as these neurons exhibit a variety of disease-associated genes, they represent an ideal platform with which to investigate neurodevelopmental mechanisms in native human cells in health and disease. Electronic supplementary material The online version of this article (doi:10.1186/s13287-015-0136-8) contains supplementary material, which is available to authorized users. Introduction In the past decade, advances in stem cell biology have led to the emergence of novel powerful tools to investigate complex questions in neurobiology. Human neural stem cells (hNSCs) and the neural progenitor cells (NPCs) that they generate have become a major focus of interest as they provide a renewable and accessible model system in which to investigate basic human neurodevelopment mechanisms and complex neurodevelopmental disorders [1C4]. One major advantage of hNPCs is usually that they can easily undergo biochemical, pharmacological and genetic manipulations, making them an ideal platform for high-throughput, genetic or small molecule functional screening [1C3, 5, 6]. Human NSCs and NPCs have been derived from numerous stem cell types, including embryonic, fetal and adult stem cells [7C10]. Previous studies have exhibited that hNPCs are self-renewing and are multipotent, being able to differentiate into multiple neural cell types, including different types of neurons, astroctyes and oligodendrocytes [11C14]. Many groups MPO-IN-28 have successfully generated neurons characteristic of different neural tissues, including spinal electric motor neurons, spinal-cord interneurons, midbrain cortical and dopaminergic pyramidal neurons, from rodent and individual embryonic stem cells [11C17]. Nevertheless, the logistical and moral factors from the usage of individual blastocytes, that embryonic stem cells are produced, makes this process challenging frequently, when investigating the essential mechanisms underlying neurodevelopment specifically. An alternative solution approach continues to be the creation of immortalised hNPCs conditionally, produced from post-mortem individual fetal tissues [2, 5, 6, 14]. Several clonal Recently, conditionally immortalised hNPC lines had MPO-IN-28 been isolated from initial trimester individual fetal tissue [14]. These cells had been conditionally immortalised using retroviral integration of an individual copy from the c-mycERTAM build. Thus, in the current presence of 4-hydroxytamoxifen (4-OHT) and described growth elements, these hNPCs retain their self-renewing properties. Nevertheless, upon drawback of trophic and 4-OHT support, as well as the addition of a medium that promotes neuronal differentiation, these cells terminally differentiate into functional neurons that MPO-IN-28 retain regional identity [14]. Indeed, immortalised hNPCs isolated from first trimester human fetal spinal cord, midbrain, hippocampus and cortex have been successfully differentiated into functional neurons and interneurons both and [12, 14, 18, 19]. These hNPC lines have already been used to investigate the mechanisms of antidepressant drug action [18], to characterise the biological functions of susceptibility genes for schizophrenia and bipolar disorder [20] and are currently in trial for engraftment following ischaemic stroke [21]. However, in order to fully understand how such mechanisms may.