Brain-specific Foxp1 deletion impairs neuronal development and causes autistic-like behaviour

Abstract

Neurodevelopmental disorders are multi-faceted and can lead to intellectual disability, autism spectrum disorder and language impairment. Mutations in the Forkhead box FOXP1 gene have been linked to all these disorders, suggesting that it may play a central role in various cognitive and social processes. To understand the role of Foxp1 in the context of neurodevelopment leading to alterations in cognition and behaviour, we generated mice with a brain-specific Foxp1 deletion (Nestin-Cre(Foxp1-/-)mice). The mutant mice were viable and allowed for the first time the analysis of pre- and postnatal neurodevelopmental phenotypes, which included a pronounced disruption of the developing striatum and more subtle alterations in the hippocampus. More detailed analysis in the CA1 region revealed abnormal neuronal morphogenesis that was associated with reduced excitability and an imbalance of excitatory to inhibitory input in CA1 hippocampal neurons in Nestin-Cre(Foxp1-/-) mice. Foxp1 ablation was also associated with various cognitive and social deficits, providing new insights into its behavioural importance.

Figure 1

Morphological defects in the developing Foxp1 KO brain. Calbindin (a) and tyrosine hydroxylase (b) immunohistochemistry showing gross morphological disruption in the striatal region. Scale bar represents 1000 μm in adult brain sections, 500 μm in all others. (c) Quantification of the striatal region as defined by area of tyrosine hydroxylase positive staining, demonstrating a significant reduction in the striatum of Foxp1 KO brains, starting at P1. At least 12 sections from at least three WT and three KO brains were quantified for each stage. (d) Pathway analysis of microarray expression studies on P1 Foxp1 KO and WT striatal tissue showing the top 11 significantly regulated pathways.

Figure 2

Nissl staining on adult brain slices showing morphological alteration in the CA1 hippocampal region of KO brains compared with WT ( × 4) (a). (b) Higher magnification images of the CA1 region ( × 20). At least four brains were examined for each genotype. Scale bars represent 500 μm.

Figure 3

Decreased excitability and increased excitatory synaptic transmission in Foxp1 KO hippocampal pyramidal neurons. (a) Sample spike trains evoked by a 300 pA somatic current injection in a control (left) and a Foxp1 KO CA1 pyramidal neuron (right). (b) mean f-i curve for control (n=10) and Foxp1 KO neurons (n=12). ***P<0.001 (ANOVA, main effect). (c) Cell capacitance (Cm), cell resistance (Rm) and resting membrane voltage (Vrest) in control and Foxp1 KO neurons. (d) Sample traces of miniature EPSCs from WT and Foxp1 KO cells recorded in voltage clamp at −70 mV. Although the miniature excitatory postsynaptic currents peak frequency was not significantly altered, the mean peak amplitude was significantly larger in Foxp1 KO neurons (e, two-tailed t-test. *P<0.05). WT, n=12/3 cells/mice; Foxp1 KO, n=10/2 cells/mice. (f) Miniature inhibitory postsynaptic current example traces from each genotype. There was no significant change in the mean miniature inhibitory postsynaptic current peak frequency and amplitude between control (n=13/2 cells/mice) and Foxp1 KO cells (n=11/2 cells/mice). (g) Data are shown as mean±s.e.m.

Figure 4

Altered behaviours in Foxp1 KO mice. (a, b) Locomotor activity as measured by distance travelled (a) and total rearing time (b). (c) Spatial short-term memory measured by object location test. (d) Analysis of non-spatial short-term memory by object discrimination test. (e) Analysis of repetitive behaviours illustrating the total frequency of jackhammer jumping, jumping and upright wall scrabbling. (f–g) Tests of social interaction in Foxp1 KO mice including social exploration (f) and social retreat (g). (h) Measurement of prepulse inhibition (PPI) of the acoustic startle reflex (ASR). In all graphs, black bars represent WT and orange bars represent KO values. Fourteen WT and 6 Foxp1 KO mice were analysed in each test, except repetitive behaviour, where 11 WT and 6 Foxp1 KO animals were analysed.