RESUMEN
Genetically modified mice by means of homologous recombination in embryonic stem (ES) cells are generated by injection of manipulated ES cells into recipient blastocysts. The injected blastocysts, following reintroduction into recipient foster mice, will produce chimeric mice in which the manipulated ES cells populate the germline and transmit the induced mutation to the offspring. Crossing of the chimeras' offspring bearing the targeted mutation in heterozygosis will ultimately produce mice homozygous for the specific genetic mutation. Here we describe the steps and procedures required to generate the chimeric mice leading to the transfer of a genetic mutation to the mouse germline.
Asunto(s)
Transferencia de Embrión , Animales , Blastocisto , Células Madre Embrionarias/citología , Femenino , Ratones , Ratones Endogámicos C57BL , Microinyecciones/métodos , MicromanipulaciónRESUMEN
Genetic deletion of a single allele of the BDNF gene affects hippocampal LTP and causes several behavioral phenotypes, including deficits in spatial learning. In the developing visual cortex, overexpression of BDNF accelerates the time course of the critical period for monocular deprivation (MD), and exogenous administration of BDNF alters the outcome of MD. We asked whether reduced levels of BDNF could affect visual cortex plasticity by studying long-term potentiation (LTP) induction and the effects of MD in heterozygous BDNF knock-out mice. We found that theta burst stimulation that induced LTP in the layer IV-III pathway of wild-type (wt) mice caused only a transient potentiation in BDNF+/- mice, and that this potentiation vanished in 25 min. In contrast, LTP elicited by stimulation of the white matter (WM), a form of LTP that can be induced only during the critical period, occurred normally in wt and BDNF+/- mice. The effects of MD during the critical period were similar in wt and BDNF+/- mice, indicating that layer IV-evoked, layer III LTP is not required for ocular dominance plasticity. We then asked whether reduction of cortical BDNF levels could prolong the critical period for MD and for the WM-evoked, layer III LTP induction. We found that in adult BDNF+/- mice, WM-evoked, layer III LTP was not inducible, and that the critical period for MD terminated normally. We conclude that deletion of one copy of the BDNF gene selectively impairs LTP of the layer IV-III pathway but does not alter ocular dominance plasticity.