In part due to the lack of patient material for study, the molecular mechanisms responsible for many central nervous system (CNS) diseases are not well understood. Fibroblasts from patients and controls can be directly reprogrammed into human induced pluripotent stem cells (hiPSC) and subsequently differentiated into disorder-specific neurons. hiPSCs represent a new strategy for studying disease; a number of recent publications have demonstrated that CNS diseases ranging from type I spinal muscular atrophy (Ebert et al., 2009), familial dystonia (Lee et al., 2009), Parkinson’s disease (PD) (Park et al., 2008; Soldner et al., 2009), amyotrophic lateral sclerosis (ALS) (Dimos et al., 2008), Rett's syndrome (Hotta et al., 2009; Marchetto et al., 2010), and schizophrenia (Brennand et al., 2011) can be modeled using hiPSC neurons.
A new method to generate patient neurons, obviating the time-consuming step of generating and validating hiPSCs has now been developed. Last year, it was demonstrated that viral expression of just three factors—ASCL1, BRN2, and MYT1Ll—is sufficient to convert adult mouse fibroblasts into functional induced neurons (iNs) in vitro (Vierbuchen et al., 2010). With the addition of a fourth factor—NeuroD—direct reprogramming of human cells to iNs has recently been demonstrated (Pang et al., 2011). From both mouse and human cells, the conversion is incredibly rapid, generating iNs capable of producing action potentials within 14 days, and permanent, with stable neuronal fate maintained up to three weeks following the repression of viral genes. Compared to mouse iNs, however, human iNs seems relatively immature: they have slightly depolarized membrane potentials, lower amplitude synaptic responses, and spontaneous synaptic activity appears more slowly, being first detectable at five to six weeks.
The ability to generate iNs from healthy and diseased patients means that this may be a new tool with which to study neurological disorders. The rapid experimental timeframe of iN generation and the theoretical potential to reprogram to specific neuronal subtypes make this an appealing experimental strategy for in vitro models of neurological disease. Current technical limitations concern efficiency and neuronal identity. While the conversion is relatively efficient, occurring at an estimated rate of 2-4 percent, whether this is sufficient for disease modeling remains to be determined. Furthermore, the ability to generate specific neuronal subtypes remains undemonstrated; the current method generates a heterogeneous mix of neurons, predominantly glutamatergic and dopaminergic, expressing markers of either forebrain or peripheral neuron patterning.
Two important issues must be considered as one contemplates modeling psychiatric disorders using iNs. We worry that bypassing neuronal differentiation and maturation will shortcut the cellular phenotype of these neurodevelopmental disorders. For example, if psychiatric disease results from abnormal synaptic maturation, iN generation may bypass the developmental window in which the disease phenotype can be observed in vitro. Second, if ASCL1, BRN2, or MYT1L contribute to the disease state, persistent overexpression might be sufficient to mask or rescue cellular phenotypes in vitro. It is not unreasonable to predict that overexpression of one or more of these key neuronal genes might affect disease initiation or progression: mutations disrupting MYT1L expression and binding have been linked to schizophrenia (Vrijenhoek et al., 2008; Riley et al., 2010), BRN2 regulates expression of a conductance calcium-activated potassium channel (KCNN3) implicated in schizophrenia (Sun et al., 2001), and ASCL1 has been linked to Parkinson’s disease (Ide et al., 2005).
The most prudent course of action would be to recapitulate the cellular and molecular phenotypes observed in patient hiPSC neurons with patient iNs. With this validation complete, we predict that many studies of psychiatric disorders using iNs will begin.
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