Neurotrophins induce release of neurotrophins by the regulated secretory pathway

Recent studies have established that neurotrophin synthesis and secretion are regulated by activity and that these factors are involved in activity-dependent processes in the nervous system. Neurotrophins also are known to induce increases in intracellular calcium, a trigger for regulated secretion. This finding raises the possibility that neurotrophins themselves may stimulate regulated secretion of neurotrophins. To address this question, we studied the release of neurotrophins from transfected PC12 cells, a widely used model for neuronal secretion and neurotrophin signal transduction. We found that neurotrophins induced the regulated secretion of brain-derived neurotrophic factor, neurotrophin-3 (NT-3), and neurotrophin-4/5. The effect of brain-derived neurotrophic factor on release of NT-3 could be abolished by REX, a p75 blocking antibody, but not by K252a, an inhibitor of neurotrophin tyrosine kinase receptor (Trk) signaling. The nerve growth factor effect on release of NT-3 could be blocked only by simultaneous application of REX and K252a, suggesting that they are mediated by TrkA as well as p75. Our data show that neurotrophins are able to induce the regulated secretion of neurotrophins and suggest a signal-transducing role for both TrkA and p75 in this process. The neurotrophin-induced release of neurotrophins may be relevant for activity-dependent processes such as synaptic plasticity and memory formation.     

Neurotrophin-4/5 and neurotrophin-3 are present within the human ovarian follicle but appear to have different paracrine/autocrine functions

Neurotrophins are a family of soluble polypeptide growth factors widely recognized for their role in the mammalian nervous system. We first reported the unique presence of one neurotrophin, brain-derived neurotrophic factor (BDNF), in the follicular fluid of the human ovarian follicle. The BDNF receptor, Trk B, was identified in mouse oocytes, and BDNF accelerated first polar body extrusion in vitro. In the present study, we examined human follicular fluid and mouse immature oocytes to determine whether another Trk B ligand, neurotrophin-4/5 (NT-4/5), is present within the ovarian follicle and if so, whether it demonstrates activity similar to that of BDNF. We also examined whether a non-Trk B neurotrophin ligand, neurotrophin-3 (NT-3), is present within the follicle and has a possible role in oocyte maturation. NT-4/5 and NT-3 were noted to be present in all human follicular fluid samples aspirated from follicles of women undergoing in vitro fertilization. NT-4/5, but not NT-3, significantly promoted mouse oocyte polar body extrusion. Trk C receptors were not noted to be present in mouse oocytes. This study demonstrates for the first time that NT-4/5 and NT-3 are present in the follicular fluid of the human ovary. These data suggest that NT-4/5, like BDNF, promotes oocyte nuclear maturation. In contrast, NT-3 does not promote oocyte maturation but may contribute to follicle-oocyte maturation by mechanisms not yet identified.     

Characterization of the neurotrophic response to acute pancreatitis

INTRODUCTION: Interesting preliminary data on changes in the neurotrophin system in various digestive diseases have recently begun to emerge. AIMS: To measure changes in messenger RNA (mRNA) levels of neurotrophins and to identify cell types expressing neurotrophins in the pancreas of rats with L-arginine-induced pancreatitis. METHODOLOGY: Rats were killed at time points from 2 hours to 4 weeks after the induction of pancreatitis, and responses were measured by assay. RESULTS: By RNase protection assay, ciliary neurotrophic factor (CNTF) mRNA expression showed a rapid response (sixfold increase over control) in the inflamed pancreas at 2 hours. The levels of mRNA expression of brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4) in the inflamed pancreas reached a peak at 1 week (2.5-fold, twofold, fourfold, and fivefold increase, respectively). By immunohistochemistry, immunoreactivity for all neurotrophins examined was observed in the islets of Langerhans in the control pancreas at all time points, but it was markedly reduced in the islets in the inflamed pancreas at 2 and 6 hours. Acinar and ductal cells, inflammatory cells, and neural elements were immunoreactive for those neurotrophins in the inflamed pancreas from 2 hours to 2 weeks. CONCLUSION: The temporal and spatial expression of neurotrophins in the course of experimental pancreatitis suggests that their upregulation is a critical component of the response of the pancreas to injury in this model.     

Reversal of spatial memory impairments in aged rats by nerve growth factor and neurotrophins 3 and 4/5 but not by brain-derived neurotrophic factor.

Aged rats, displaying impairments in spatial learning and memory associated with marked cellular atrophy of forebrain cholinergic neurons, received intracerebroventricular infusions of one of the four neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), or neurotrophin 4/5 (NT-4/5), or a combination of NGF and BDNF, or vehicle. During the 4-week infusion period rats receiving NGF, NT-3, or NT-4/5 showed improved acquisition and retention of spatial memory. With NGF and NT-3, but not NT-4/5, this was accompanied by a significant reduction in cholinergic neuron atrophy in septum, nucleus basalis, and striatum. BDNF, in contrast, was without effect either alone or in combination with NGF. These results show that memory deficits associated with aging can be reversed by several members of the neurotrophin family and that this effect may be mediated through activation of multiple neurotrophin receptors associated with cholinergic and possibly noncholinergic systems in the brain.     

Neurotrophin 3 rescues neuronal precursors from apoptosis and promotes neuronal differentiation in the embryonic metanephric kidney.

We analyzed the developmental regulation and role of the neurotrophins during metanephric kidney morphogenesis. RNase protection assay revealed the presence of nerve growth factor, neurotrophin 3 (NT-3), and brain-derived neurotrophic factor mRNAs and the regulation of their expression during embryonic development of rat metanephros. NT-3 induced differentiation (neurite outgrowth) and survival (inhibition of apoptosis) of the neuronal precursors in cultured nephrogenic mesenchymes and neuronal differentiation in cultured whole kidneys, whereas NT-4/5, brain-derived neurotrophic factor, and nerve growth factor were without effect. The neurotrophins did not trigger tubular differentiation of isolated nephrogenic cells, which underwent apoptosis when cultured with or without the neurotrophins. NT-3 is thus an inducer of differentiation and a survival factor for renal neuronal cells, but none of the neurotrophins is a morphogen in kidney tubule induction.     

Rapid stimulatory effects of brain-derived neurotrophic factor and neurotrophin-3 on somatostatin release and intracellular calcium rise in primary hypothalamic cell cultures.

Although the long-lasting effects of neurotrophins have been extensively studied, less data are available on their rapid effects, especially on peptide release. In the present report, we investigated rapid effects of neurotrophins on somatostatin release and on intracellular calcium concentration ([Ca(2+)](i)) in primary cultures of hypothalamic neurons. RT-PCR experiments revealed mRNA expression of the three high-affinity neurotrophin receptors tyrosine kinase (Trk) TrkA, TrkB and TrkC, indicating potential responses to their preferential ligands: nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin 3 (NT-3), respectively. We demonstrated that BDNF, and to a lesser extent NT-3, induced significant time- and concentration-dependent somatostatin release, while NGF was devoid of any effect. BDNF or NT-3 induction of somatostatin release was inhibited by the Trk inhibitors K-252a and genistein, whereas K-252b, a less effective inhibitor, had no effect. BDNF- and NT-3-induced somatostatin release depended upon extra- and intracellular Ca(2+) since it was completely abolished in the presence of the Ca(2+) chelators BAPTA (bis-(alpha-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid) or BAPTA-AM (bis-(alpha-aminophenoxy)-ethane-N,N,N',N'-tetraacetoxymethylester), respectively. In addition, BDNF and NT-3 induced a sustained and rapid increase in [Ca(2+)](i) which depended on the extracellular Ca(2+) concentration. MK-801 (dizocilpine) and tetrodotoxin (TTX) entirely blocked neurotrophin-evoked somatostatin release and [Ca(2+)](i) rise in response to BDNF and NT-3 application in most neurons. Neurotrophin-induced [Ca(2+)](i) rise was completely blocked by K-252a. The present results are consistent with: (1) an indirect effect of neurotrophins on somatostatin release via endogenous glutamate release and subsequent NMDA receptor activation, (2) a major indirect effect of neurotrophins on Ca(2+) rise in hypothalamic neurons which very likely occurs through NMDA receptor activation. Taken altogether, these results indicate that BDNF and NT-3 can rapidly affect the activity of hypothalamic neurons.     

Activation of calcium-dependent potassium channels in rat brain neurons by neurotrophin-3 and nerve growth factor

The neurotrophins are signaling factors that are essential for survival and differentiation of distinct neuronal populations during the development and regeneration of the nervous system. The long-term effects of neurotrophins have been studied in detail, but little is known about their acute effects on neuronal activity. Here we use permeabilized whole-cell patch clamp to demonstrate that neurotrophin-3 (NT-3) and nerve growth factor activate calcium-dependent, paxilline-sensitive potassium channels (BK channels) in cortical neurons. Application of NT-3 or nerve growth factor produced a rapid and gradual rise in BK current that was sustained for 30–50 min; brain-derived neurotrophic factor, ciliary neurotrophic factor, and insulin-like growth factor-1 had no significant effect. The response to NT-3 was blocked by inhibitors of protein kinases, phospholipase C, and serine/threonine protein phosphatase 1 and 2a. Omission of Ca²⁺ from the extracellular medium prevented the NT-3 effect. Our results indicate that NT-3 stimulates BK channel activity in cortical neurons through a signaling pathway that involves Trk tyrosine kinase, phospholipase C, and protein dephosphorylation and is calcium-dependent. Activation of BK channels may be a major mechanism by which neurotrophins acutely regulate neuronal activity.