Distribution of mRNAs in cultured thalamic cells, explants, and their axons. (A) Photomicrograph of cultured thalamic explant (on left) with axons growing from it stained with an antibody against neurofilament. Broken lines outline the regions cut out to obtain samples for RNA extraction. (B) A portion of a gel showing PCR amplified products from mRNAs taken from area 1 'Cells' (left lane) and area 2 'Axons' (right lane). (C) Axon:Cell ratios for 8 mRNAs measured by qRT-PCR. Each point shows mean ± SEM for 3 samples. (D) in situ hybridisation for β-catenin mRNA (black signal) on a dissociated thalamic neuron. (E-G) Quantification of hybridisation signal density for (E) β-catenin mRNA, (F) β-actin mRNA, and (G) 18S rRNA with antisense probe signal in red and background signal in green. Lines used for densitometric analysis were drawn along the axon connecting the cell body (cb- 0% along the line) and growth cone (gc- 100% along the line) as indicated in (D). The β-catenin antisense probe (n = 12) and β-actin antisense probe (n = 11) are compared against a β-catenin sense probe (n = 15), whereas the 18S oligoprobe (n = 6) is compared against a Scrambled oligoprobe (n = 7). The sense and scrambled control probes are used to define background signal levels. An asterisk above a data point indicates significant difference from background (Student's t-test, p < 0.05). Note that each transcript has a distinct distribution profile along the axon. Abbreviations: cell body (cb); growth cone (gc); axon (a). Scale bars: A, 100 μm; E, 10 μm.
Pratt et al. BMC Neuroscience 2012 13:20 doi:10.1186/1471-2202-13-20