Figure 4.

DsRed and tau-1 are colocalized and branch numbers are increased in hippocampal cells in the absence of FAK at 6 DIV in vitro. At early stages, most of the neurites in these cultures express tau-1 in neurons transfected with control (shown here) and Cre- or FRNK-expressing plasmids (not shown). (a,b,d,f) Expression of DsRed and pCDNA. (c,e,g) Tau-1 immunoreactivity. (b,d,f) Higher magnification images corresponding to the boxed regions indicated in a. Most branching neurites at these stages are developing axons as they express the axon-specific marker tau-1 (∼95%, n = 10 neurons). Arrows help to position the colocalization. Filled arrows indicate axons (tau-positive neurites), open arrows indicate dendrites (tau-negative neurites), white asterisks indicate branchpoints. (h–p) fak^flox deletion in mouse hippocampal cultures. (h–m) Fluorescence images and representative drawings of neurons transfected with control pCDNA3 (h–j) or Cre plasmid (k–m) at 6 DIV. (n) Quantification of branches at increasing branch orders per neuron. *P < 0.01 to P < 0.0001 versus control by two-tailed t-test (n = 92 neurons from four cultures in n–p). (o) Quantification of the total neurite length per neuron expressed in µm.*P < 0.0005 versus control (control 1,241.36 ± 93.24, mutant 2,018.04 ± 169.44). (p) Quantification of the total branches per µm of axon. *P < 0.0005 versus control (control 0.0045 ± 0.0003, mutant 0.0067 ± 0.0005). Note that, as described above, most branching neurites at these stages are developing axons (∼95%), although a few of them could be dendrites. Data are the mean ± s.e.m. Scale bars, 200 µm (n,m); 100 µm (a); 10 µm (b–g).