A. Balkowiec et Dm. Katz, Activity-dependent release of endogenous brain-derived neurotrophic factorfrom primary sensory neurons detected by ELISA in situ, J NEUROSC, 20(19), 2000, pp. 7417-7423
To define activity-dependent release of endogenous brain-derived neurotroph
ic factor (BDNF), we developed an in vitro model using primary sensory neur
ons and a modified ELISA, termed ELISA in situ. Dissociate cultures of nodo
se-petrosal ganglion cells from newborn rats were grown in wells precoated
with anti-BDNF antibody to capture released BDNF, which was subsequently de
tected using conventional ELISA. Conventional ELISA alone was unable to det
ect any increase in BDNF concentration above control values following chron
ic depolarization with 40 mM KCl for 72 hr. However, ELISA in situ demonstr
ated a highly significant increase in BDNF release, from 65 pg/ml in contro
l to 228 pg/ml in KCl-treated cultures. The efficacy of the in situ assay a
ppears to be related primarily to rapid capture of released BDNF that preve
nts BDNF binding to the cultured cells. We therefore used this approach to
compare BDNF release from cultures exposed for 30 min to either continuous
depolarization with elevated KCl or patterned electrical field stimulation
(50 biphasic rectangular pulses of 25 msec, at 20 Hz, every 5 sec). Short-t
erm KCl depolarization was completely ineffective at evoking any detectable
release of BDNF, whereas patterned electrical stimulation increased extrac
ellular BDNF levels by 20-fold. In addition, the magnitude of BDNF release
was dependent on stimulus pattern, with high-frequency bursts being most ef
fective. These data indicate that the optimal stimulus profile for BDNF rel
ease resembles that of other neuroactive peptides. Moreover, our findings d
emonstrate that BDNF release can encode temporal features of presynaptic ne
uronal activity.