Homozygous mutation of focal adhesion kinase in embryonic stem cell derived neurons: normal electrophysiological and morphological properties in vitro

Mouse ES cells carrying homozygous null mutations (FAK-/-) were generated and differentiated in vitro into neurons. FAK-/- ESNs extended axons and dendrites and formed morphologically and electrophysiologically intact synapses. A detailed study of NMDA receptor gated currents and voltage sensitive calcium currents revealed no difference in their magnitude, or modulation by tyrosine kinases.FAK does not have an obligatory role in neuronal differentiation, synapse formation or the expression of NMDA receptor or voltage-gated calcium currents under the conditions used in this study. The use of genetically modified ESNs has great potential for rapidly and effectively examining the consequences of neuronal gene manipulation and is complementary to mouse studies.A major goal in post-genomic neuroscience is to elucidate the function of individual genes in neurons. Gene targeting is one of the most widely used methods to study the function of genes in the vertebrate nervous system and in recent years the need for conditional mutation strategies has been emphasised. For example, the tetracycline system has been used to transiently express genes in the brain [1] and the Cre/LoxP system used to delete genes in selected neuronal populations [2]. One disadvantage to these techniques is that it can take 2 or more years for results, in part because of the breeding of mice. Therefore any method that allows for a shorter duration and reduces the need for animals is potentially useful. Toward this end, we have adopted an experimental strategy using genetically engineered embryonic stem (ES) cell derived neurons (ESNs). This system has the potential to greatly speed the functional analysis of genetic manipulations and importantly also allows access to lethal mouse phenotypes.Totipotent murine ES cells can be induced to differentiate in vitro to form mature post-mitotic neurons, which in addition to being synaptically connected, show many properties comparable to primary cultures of em