Fig. 3. Diminished innervation of the skin in K14-BMP4 mice. Innervation of the epidermis and upper dermis at comparable locations in the intervibrissal fur in wild type and K14-BMP4 mice. Labels are similar to those in Fig. 2. (A-D) Double-label immunofluorescence with anti-CGRP (Cy3) and anti-PGP (Cy2). At 1 month, the innervation to the epidermis appears comparable in wild type and K14-BMP4 mice. At 6 months, innervation in the K14-BMP4 mice has become reduced and fragmented. However, most of the remaining innervation expresses CGRP-IR, which may be more than in wild type. Yellow arrows indicate sites where axons express CGRP IR. In contrast to wild type, in K14-BMP4 transgenics most epidermal innervation seems to be supplied directly from the third tier of the dermal plexus. Axon bundles at the epidermal-dermal border, indicative of the fourth tier, are readily encountered in wild type. The fourth tier was rarely detected in K14-BMP4 intervibrissal fur. A-D are all mice from CB6F1 background. (E-H) Comparison of the innervation to the epidermis and upper dermis at comparable locations in the intervibrissal fur and in wild-type, K14-BMP4 and K14-noggin mice as shown by double-label immunofluorescence with anti-CGRP (Cy3) and anti-NF200 (Cy2). E,F are from a CB6F1 background; G,H are from a Fvb background. Red arrows indicate axons labeled only with anti-CGRP, green arrows labeled only with anti-NF200, yellow arrows labeled with both anti-CGRP and anti-NF. Note the massive increase in CGRP-positive innervation at all levels of the dermal plexus in the K14-noggin fur. In both wild-type and K14-noggin mice, the third and fourth tiers of the dermal plexus contain anti-CGRP labeled axons that are mostly NF200 negative (red arrows). Likewise, the anti-CGRP labeled endings in the epidermis lack NF200-IR (red arrowheads). A few axons in the third tier contain NF200 but most of the CGRP-positive/NF200-positive axons are in the second tier. By contrast, in the K14-BMP4 fur, most of the CGRP positive innervation at all levels co-expresses NF200-IR. This includes an ending in the epidermis that co-expresses CGRP-IR and NF200-IR (yellow arrowhead), which is never seen in the wild-type epidermis.