Target‐derived BDNF (brain‐derived neurotrophic factor) is essential for the survival of developing neurons in the isthmo‐optic nucleus

Abstract

Neurons in the peripheral nervous system depend on single neurotrophic factors, whereas those in the brain are thought to utilize many different trophic factors. This study examined whether some neurons in the brain critically depend on a single trophic factor during development. Neurons in the isthmo-optic nucleus (ION) of chick embryos respond to exogenous brain-derived neurotrophic factor (BDNF). Relatively high concentrations of endogenous BDNF were present in the ION of 14-18-day-old chick embryos. ION target cells in the retina were immunolabeled for BDNF but showed surprisingly low levels of BDNF mRNA. These data suggest that ION target cells derive some BDNF from other retinal sources. No BDNF mRNA was detected in the ION itself. ION neurons had a very efficient retrograde transport system for BDNF and exogenous BDNF arrived in the ION intact. When the ION was deprived of endogenous trkB ligands by injection of trkB fusion proteins in the eye, cell death of ION neurons was enhanced, and this effect was mimicked by BDNF-specific blocking antibodies in the eye. TrkB fusion proteins in the retina induced cell death of ION neurons prior to visible effects on ION target cells in the retina. Immunolabel for endogenous BDNF was sparse in pyknotic ION neurons, suggesting that ION neurons with low BDNF content were eliminated by apoptosis. These data show that BDNF is an essential target-derived trophic factor for developing ION neurons and thereby validate the neurotrophic hypothesis for at least one neuronal population in the brain.

Presynaptic neurotrophin‐3 increases the number of tectal synapses, vesicle density, and number of docked vesicles in chick embryos

“…Two of the examined parameters, number of synapses/area and density of vesicles/synapse area, decreased with deprivation of endogenous TrkB-ligands, whereas only one parameter, vesicle density, was increased with increased levels of BDNF. This finding suggests that either endogenous NT-4 may have functions on synapse numbers, in addition to BDNF, and/or that endogenous BDNF levels in the tectum are already at maximal functional levels, consistent with reports of relatively high BDNF expression and protein levels in the chick optic tectum compared with other neuronal tissues such as retina (Herzog et al, 1994;Johnson et al, 1997;Herzog and von Bartheld, 1998;von Bartheld and Johnson, 2001). Therefore, increased BDNF levels may fail to further increase parameters of synaptic strength.…”

Fates of Neurotrophins after Retrograde Axonal Transport: Phosphorylation of p75NTR Is a Sorting Signal for Delayed Degradation

“…The amounts of radioactivity in the dissected eyes and isthmo-optic nuclei (IONs) were measured in a gamma-counter (Wallac 1470 Wizard). Radioactivity visualized as silver grains in ION neurons likely represents intact neurotrophin, because a significant fraction of radioactivity recovered from the ION after retrograde axonal transport is intact protein as shown by SDS PAGE (von Bartheld and Johnson, 2001). …”

“…The brains were postfixed and the midbrains and brainstems were serially sectioned in the transverse plane at 500 µm on a vibratome (series 1000). From these slices, the left and right IONs were dissected as described (von Bartheld and Johnson, 2001), and each ION slice was gamma-counted. The cpm amounts of the slices containing parts of the ION were added.…”

“…The ION was labeled retrogradely by intraocular injection of the lipophilic dye DiI (1–2% in 100% ethanol, Molecular Probes) and dissected under an inverted fluorescence microscope (von Bartheld and Johnson, 2001). For each individual experiment the total RNA from two to three IONs was extracted.…”

“…The animals were killed at ages E14-15 by decapitation, and heads were fixed by immersion in Methacarn’s fixative. The brains were dissected, dehydrated in methanol, and embedded in paraffin as previously described (von Bartheld et al, 1996a; Janiga et al, 2000; von Bartheld and Johnson, 2001). The isthmic region of the brain containing the right and left IONs was serially sectioned in the transverse plane at 12 µm.…”