Abundant production of brain-derived neurotrophic factor by adult visceral epithelia. Implications for paracrine and target-derived Neurotrophic functions

Abstract

Brain-derived neurotrophic factor (BDNF) plays a crucial role for the survival of visceral sensory neurons during development. However, the physiological sources and the function of BDNF in the adult viscera are poorly described. We have investigated the cellular sources and the potential role of BDNF in adult murine viscera. We found markedly different amounts of BDNF protein in different organs. Surprisingly, BDNF levels in the urinary bladder, lung, and colon were higher than those found in the brain or skin. In situ hybridization experiments revealed that BDNF mRNA was made by visceral epithelial cells, several types of smooth muscle, and neurons of the myenteric plexus. Epithelia that expressed BDNF lacked both the high- and low-affinity receptors for BDNF, trkB and p75(NTR). In contrast, both receptors were present on neurons of the peripheral nervous system. Studies with BDNF-/-mice demonstrated that epithelial and smooth muscle cells developed normally in the absence of BDNF. These data provide evidence that visceral epithelia are a major source, but not a target, of BDNF in the adult viscera. The abundance of BDNF protein in certain internal organs suggests that this neurotrophin may regulate the function of adult visceral sensory and motor neurons.

Figures

Figure 1.

BDNF protein concentrations in internal organ lysates. BDNF protein concentrations in total organ lysates of 8-week-old Balb/c mice were measured by ELISA and calculated as nanograms of BDNF per gram of total protein. Shown are the means with each standard deviation (n = 4). A: BDNF protein in the total brain, back skin and cardiorespiratory system. B: BDNF protein in the gastrointestinal tract. C: BDNF protein in the urogenital tract.

Figure 2.

BDNF expression in internal organs. Detection of BDNF mRNA was performed on 10-μm cryosections by ISH. Sequential sections were hybridized with anti-sense or sense riboprobes, respectively. Shown are the cortex of the kidney, a liver lobe with central vein, mucosa and tunica muscularis of the stomach, and sigmoid colon. Note the BDNF mRNA negative glomerula in the kidney. The arrow in the micrograph of the colon shows a BDNF mRNA-positive ganglion of the myenteric plexus. Scale bar, 27 μm. L, lumen; CV, central vein; G, glomerulum; SM, smooth muscle; BM, basal membrane and submucosa of the stomach.

Figure 3.

BDNF expression in visceral epithelia. Sequential cryosections were hybridized with anti-sense and sense riboprobes. The first slide of each set was stained with hematoxylineosine (HE). Shown are the mucosa and tunica muscularis of the upper part of esophagus, of a medium-sized bronchus in the lung, of the urinary bladder, the squamous epithelium on the portio vaginalis uteri, mucosa and submucosa of the oviduct, and the peritoneal serosa layer on the myometrium muscle. Arrows indicate the basal membrane of the epithelia. Note that also adjacent stroma cells in the oviduct appear BDNF mRNA-positive. On the negative myometrium background, peritoneal BDNF mRNA signals are easily to identify by blue staining in the peritoneum. Scale bar, 13.5 μm. L, lumen; A, airway; C, peritoneal cavity.

Figure 4.

BDNF receptors in the colon and cervix uteri. Immunohistochemistry was performed against the full-length trkB and p75NTR receptors on 10-μm cryosections of Balb/c mice internal organs. Sections were counterstained by Hoechst 33342 (blue fluorescence) for identification of cell nuclei (trkB) or by haemalaun (p75NTR). Incubation of inner organ cryosections without primary antibody served as a negative control (not shown). trkB (red fluorescence) or p75NTR staining (red APAAP staining) was evaluated in comparison to this background. Shown are sections of the portio vaginalis uteri and transverse colon. Arrowheads in all figures indicate epithelia, arrows in the micrograph of colon neurons and nerve fibers of the myenteric plexus. Scale bar, 50 μm.

Figure 5.

Epithelia of BDNF−/− mice. Two-micron paraffin sections of 2-week-old wild-type and BDNF−/− mice internal organs were HE-stained following standard laboratory procedures. Shown are sections of the transverse colon wall and respiratory epithelium and airway smooth muscle of a main bronchus of wild-type and BDNF−/− mice. Note the mucosal atrophy and enlarged goblet cells (GC) in the BDNF−/− colon. Lungs were indistinguishable between wild-type and BDNF−/− mice; the respiratory epithelium was unaltered in morphology and height. Scale bar, 13.5 μm. L, lumen; A, airway; GC, goblet cells; SM, smooth muscle.