Region-specific neurotrophin imbalances in Alzheimer disease: decreased levels of brain-derived neurotrophic factor and increased levels of nerve growth factor in hippocampus and cortical areas

BACKGROUND: Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), and neurotrophin 4/5 (NT-4/5) are members of the neurotrophin gene family that support the survival of specific neuronal populations, including those that are affected by neurodegeneration in Alzheimer disease (AD). OBJECTIVE: To determine whether neurotrophin protein levels are altered in the AD-affected brain compared with control brains. METHODS: We quantitated protein levels of NGF, BDNF, NT-3, and NT-4/5, and calculated neurotrophin/NT-3 ratios in AD-affected postmortem hippocampus, frontal and parietal cortex, and cerebellum, and compared them with age-matched control tissue (patients with AD/controls: hippocampus, 9/9 cases; frontal cortex, 19/9; parietal cortex, 8/5; and cerebellum, 5/7, respectively). We applied highly sensitive and specific enzyme-linked immunosorbent assays in rapid-autopsy-derived brain tissue (mean+/-SD postmortem interval, 2. 57+/-1.75 h, n=71) to minimize postmortem proteolytic activity. RESULTS: Levels of BDNF were significantly reduced in hippocampus and parietal cortex (P<.001, and P<.01) as well as BDNF/NT-3 ratios in frontal and parietal cortices (P<.05, and P<.01) in the group with AD compared with the control group. Levels of NGF and NGF/NT-3 ratio were significantly elevated in the group with AD compared with the control group in the hippocampus and frontal cortex (P<.001). Levels of NT-4/5 and the NT-4/NT-3 ratio were slightly reduced in hippocampus and cerebellum in the group with AD compared with the control group (P<.05). In contrast, the levels of NT-3 were unchanged in all brain regions investigated. CONCLUSION: Decreased levels of BDNF may constitute a lack of trophic support and, thus, may contribute to the degeneration of specific neuronal populations in the AD-affected brain, including the basal forebrain cholinergic system. Arch Neurol. 2000.     

Thyroid hormone regulation of NGF, NT-3 and BDNF RNA in the adult rat brain.

The effects of peripherally administered thyroid hormone (TH; 500 micrograms/kg; i.p.; q.d.) on the relative abundances of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) RNA were determined by rtPCR in the cortex and hippocampus of young adult rats. Corresponding changes in choline acetyltransferase (ChAT) activity were measured since NGF and BDNF have been shown to enhance the expression of this marker enzyme of central cholinergic pathways. Abundance levels of NGF and NT-3, relative to cyclophilin (cycl), were increased significantly (+50%, P < 0.05) in the hippocampus following TH treatment. Despite enhanced abundance of NGF in the hippocampus, ChAT activity was unchanged, whereas ChAT activity was modestly increased by 28% in the cortex without corresponding changes in NGF, NT-3 or BDNF. These results demonstrate that TH administration is capable of inducing the accumulation of NT-3, in addition to NGF but that the induction levels of RNA cannot be directly correlated with responsivity of the cholinergic system as measured by ChAT activity.     

Regulation of Neurotrophin mRNA Expression in the Rat Brain by Glucocorticoids

Northern blot analysis was used to examine the effects of glucocorticoids on neurotrophin mRNA expression in the rat cerebral cortex and hippocampus. The results show that 3 days after adrenalectomy the mRNA levels for nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) decreased significantly in both these regions. In adrenalectomized animals given dexamethasone replacement the mRNA levels for the three neurotrophins were restored to control levels. The effect of a single dose of dexamethasone (5 mg/kg) administered i p. to intact animals on the expression of neurotrophins was also examined. NGF and NT-3 mRNAs showed a 2.5-fold and a 1.4-fold increase, respectively, during the first 4 h after the injection. The increase was followed by a decrease, with levels -50% of control 24 and 48 h after the injection. In contrast, the level of BDNF mRNA did not change during the first 10 h after the injection, but decreased to 70% of control 48 h after the injection. These data indicate that glucocorticoids regulate neurotrophin mRNA expression both in the cortex and in the hippocampus, and suggest further that the known effects of glucocorticoids on neuronal survival in the brain could be due to changes in the levels of neurotrophins in the brain.     

Electroconvulsive stimuli alter nerve growth factor but not brain-derived neurotrophic factor concentrations in brains of a rat model of depression

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are proteins involved in neuronal survival and plasticity of dopaminergic, cholinergic and serotonergic neurons in the central nervous system (CNS). Moreover, it has been hypothesized that these molecules play a role in the pathophysiology as well as treatment of depression. Using an animal model of depression, the Flinders Sensitive Line (FSL) rats and their controls, the Flinders Resistant Line (FRL), we investigated the effects of electroconvulsive stimuli (ECS) on brain NGF and BDNF. ECS or SHAM ECS were administered eight times, with a 48-h interval between each treatment. NGF and BDNF were measured with enzyme-linked immunosorbent assay (ELISA). In the hippocampus ECS increased NGF concentration in FSL but not FRL rats. ECS decreased NGF concentration in the frontal cortex of FSL rats. In both FSL and FRL rats ECS increased NGF levels in the striatum. In contrast, ECS did not change BDNF concentration in hippocampus, frontal cortex and striatum of FSL and FRL rats. Our data support the notion that neurotrophin concentrations may be altered by ECS.     

Electroconvulsive stimuli alter the regional concentrations of nerve growth factor,brain-derived neurotrophic factor,and glial cell line-derived neurotrophic factor in adult rat brain

In this study we investigated whether electroconvulsive stimuli (ECS) altered the regional brain protein concentrations of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and glial cell line-derived neurotrophic factor (GDNF) in Sprague Dawley rats. Electroconvulsive stimuli were administered once daily for 8 days. At the end of the experiment, rats were killed, the brains were dissected into five regions, and the neurotrophic factors were extracted and measured by enzyme-linked immunosorbent assay. Electroconvulsive stimuli increased the concentrations of NGF in the frontal cortex and concentrations of BDNF in the hippocampus, the striatum, and the occipital cortex. In contrast, ECS decreased GDNF concentrations in the hippocampus and the striatum. Our data indicate that neurotrophic factors play a role in the mechanism of action of ECS and, by extrapolation, may play a role in the mechanism of action of electroconvulsive treatment.     

Age-dependent time course of cerebral brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3 in APP23 transgenic mice

Neurotrophins, including brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3), have repeatedly been shown to be involved in the pathophysiology of Alzheimer's disease (AD). Recent studies have claimed that these neurotrophic factors are important tools for therapeutic intervention in neurodegenerative diseases. So far, little is known about the age- and disease-modulated time course of cerebral neurotrophins. Therefore, we have studied protein concentrations of BDNF, NGF, and NT-3 in different brain areas and sciatic nerve, a neurotrophin-transporting peripheral nerve, in a well-characterized AD model of amyloid precursor protein-overexpressing rodents (APP23 mice) at the ages of 5.0, 10.5, and 20.0 months. In APP23 mice, there was a significant increase of BDNF and NGF in the frontal and occipital cortices (for BDNF also in the striatum) of old 20.0-month-old mice (with respect to median values up to 8.2-fold), which was highly correlated with amyloid concentrations of these brain areas. Median values of NGF and NT-3 showed up to a 6.0-fold age-dependent increase in the septum that was not detectable in APP23 mice. Hippocampus, olfactory bulb, and cerebellum (except NT-3) did not show substantial age- or genotype-related regulation of neurotrophins. In the sciatic nerve, BDNF and NGF levels are increased in5-month-old APP23 mice and decrease with age to control levels. In conclusion, APP23 mice show a genotype-dependent increase of cortical BDNF and NGF that is highly correlated with amyloid concentrations and may reflect an amyloid-related glia-derived neurotrophin secretion or an altered axonal transport of these neurotrophic factors.     

Differential regulation of nerve growth factor and brain-derived neurotrophic factor in a mouse model of learned helplessness

Stress-induced helplessness in rodents constitutes a well-defined model to investigate neurobiological mechanisms of depression. Neurotrophins like nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) have both been shown to be involved in neurobiological changes of physiological and pathological reactions to stress. In this study we investigated NGF and BDNF protein levels in the frontal cortex and hippocampus in mice treated with an established model of inducible helplessness via electric footshocks compared to untreated controls at various times (0 h up to 14 days after treatment). NGF levels were transiently decreased by one forth in the frontal cortex of shocked mice at 6 h after the stress treatment, whereas BDNF levels remained unchanged in the brain areas investigated throughout the time course. In addition, frontal cortex BDNF levels showed a significantly higher concentration in the right compared to the left hemisphere (up to 3-fold). This effect was detectable independently of treatment, namely in shocked and control mice at any time point measured. In conclusion, a transient decrease of frontal NGF constitutes the most striking correlate of neurobiological changes in this animal model of stress-induced change of behaviour. Interhemispherical differences of BDNF content in the frontal cortex are a new finding that might reflect intracerebral side dominance. Thus, subsequent studies of frontal cortex BDNF expression should carefully consider an interhemispherical variance to avoid misinterpretation.     

Ethanol-induced alterations in the expression of neurotrophic factors in the developing rat central nervous system

Neonatal rats were exposed to ethanol throughout gestation, or during the early postnatal period (postnatal days 4-10 (P4-10)), and enzyme-linked immunoabsorbent assays were subsequently conducted in order to assess nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) protein content in hippocampus, septum, cortex/striatum and cerebellum. These determinations revealed that following prenatal ethanol treatment, there were significant ethanol-induced increases in NGF in P1 cortex/striatum, but no changes in any of the three neurotrophic factors (NTFs) in the other brain regions. Cortex/striatal NGF protein returned to control levels by P10. Following early postnatal exposure, BDNF was elevated in hippocampus and cortex/striatum (assessed on P10), and NGF was also enhanced in cortex/striatum at this age. Hippocampal and cortex/striatal BDNF returned to control levels by P21, but cortex/striatal NGF levels remained enhanced at this age. This NTF did not differ in ethanol and control animals by P60, however. The possible significance of elevated levels of NTFs as a function of ethanol exposure is discussed, and it is speculated that while such alterations could play a protective role, increases in these substances during critical developmental periods could also prove to be deleterious, and could even contribute to certain of the neuropathologies which have been observed following developmental ethanol exposure.     

Comparison of the Temporary Dynamics of NGF and BDNF Gene Expression in Rat Hippocampus,Frontal Cortex,and Retina Under Semax Action

Neurotrophins are a family of structurally related proteins that regulate the survival, differentiation, and maintenance of function of different neuron populations. Some peptides are able to affect the production and activity of neurotrophins. One of these synthetic peptides is heptapeptide Semax, an analog of the N-terminal adrenocorticotropic hormone fragment 4-10. It is known that Semax has effects on learning and memory formation and exerts some neuroprotective effects in rodents and humans. Male Wistar rats were treated for 20 min, 40 min, 90 min, 3 h, 8 h, and 24 h with Semax. Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) gene expression in rat brain and retina was analyzed by real-time polymerase chain reaction. It was revealed that after Semax administration the multidirectional activation of the expression of the genes under investigation in the hippocampus, frontal cortex, and retina was observed. The expression of both neurotrophin genes was decreased in rat hippocampus and retina 20 min after Semax administration and was increased in the frontal cortex. The expression levels of NGF remained practically constant in the retina at the initial stage, whereas the expression levels of BDNF were significantly increased 90 min after Semax administration.     

Hippocampal neurotrophin levels after injury: Relationship to the age of the hippocampus at the time of injury

Aging impairs the competence of the hippocampus for synaptic reorganization after injury. This potentially is due to the inability of the aging hippocampus to up-regulate the critical neurotrophic factors for prolonged periods after injury to levels at which they can stimulate neurite outgrowth and facilitate synaptic reorganization. We hypothesize that the concentrations of neurotrophins in the hippocampus after injury depend on the age at the time of injury. We quantified the concentrations of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3) in the hippocampus of young, middle-aged, and aged Fischer 344 rats at 4 days after kainic acid (KA)-induced injury. In comparison with the age-matched intact hippocampus, the KA-lesioned hippocampus exhibited increased levels of BDNF and NGF in all three age groups. In contrast, the NT-3 concentration was unaltered after KA lesion. Notwithstanding similar percentage increases in BDNF after injury, the lesioned middle-aged and aged hippocampus contained 45-52% less BDNF than the lesioned young hippocampus. NGF and NT-3 levels after injury were comparable across the three age groups, however. Furthermore, lower BDNF concentration in the injured aging hippocampus was associated with normal astrocytic response but significantly diminished microglial reaction. Thus, in comparison with the injured young hippocampus, the injured aging hippocampus contains considerably less BDNF but similar levels of NGF and NT-3. Lower BDNF levels in the injured aging hippocampus might underlie the diminished spontaneous healing response observed in the aging hippocampus after injury, particularly in terms of synaptic reorganization and dentate neurogenesis.     

Stress-resistant mice overexpressing glucocorticoid receptors display enhanced BDNF in the amygdala and hippocampus with unchanged NGF and serotonergic function

Dysfunctional glucocorticoid receptor (GR) signaling has been shown to be involved in the pathogenesis of depressive behavior in mice and humans. In accordance with this hypothesis GR overexpressing mice are less susceptible to develop depressive-like behavior when subjected to stressful events. Here, we analyzed GR overexpressing mice for morning and evening content of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and the tissue levels of serotonin and its metabolite 5-hydroxyindoleacetic acid) in brain areas suspected to be involved in stress adaptation. BDNF concentrations in the hippocampus and amygdala/piriform cortex were significantly enhanced in GR overexpressing mice (by maximally +103%) compared to wildtype animals. Diurnal variations, as detected for NGF in the hypothalamus, for BDNF in the frontal cortex and striatum and for serotonergic function in the frontal cortex and hypothalamus, were not affected by the genotype. In conclusion, GR overexpression-dependent increases of hippocampal and amygdala BDNF content presumably represent a dynamic correlate of enhanced stress resistance.     

Effects of electroconvulsive seizures and antidepressant drugs on brain-derived neurotrophic factor protein in rat brain.

BACKGROUND: The antidepressant-like effects of brain-derived neurotrophic factor (BDNF) infusions in brain, and the upregulation of BDNF mRNA and its receptor in rats exposed to electroconvulsive seizure (ECS) and antidepressants, suggested a role for increased BDNF protein. METHODS: We measured BDNF protein levels with a two-site enzyme-linked immunosorbent assay (ELISA) in six brain regions of adult male rats that received daily ECS or daily injections of antidepressant drugs. RESULTS: The BDNF ELISA method was validated by the 50% loss of BDNF protein in the brains of +/- BDNF knockout mice, the 60%-100% recovery of spiked recombinant BDNF, and by the amounts and regional variations of BDNF measured in the six brain regions. Ten consecutive daily exposures to ECS increased BDNF protein in the parietal cortex (219%), entorhinal cortex (153%), hippocampus (132%), frontal cortex (94%), neostriatum (67%), and septum (29%). BDNF increased gradually in the hippocampus and frontal cortex, with a peak response by the fourth day of ECS. Increases peaked at 15 hours after the last ECS and lasted at least 3 days thereafter. Two weeks of daily injections with the monoamine (MAO)-A and -B inhibitor tranylcypromine (8-10 mg/kg, IP) increased BDNF by 15% in the frontal cortex, and 3 weeks treatment increased it by 18% in the frontal cortex and by 29% in the neostriatum. Tranylcypromine, fluoxetine, and desmethylimipramine did not elevate BDNF in the hippocampus. CONCLUSIONS: Elevations in BDNF protein in brain are consistent with the greater treatment efficacy of ECS and MAO inhibitors in drug-resistant major depressive disorder and may be predictive for the antidepressant action of the more highly efficacious interventions.     

Expression of the mRNA of neurotrophins in brain regions of rats after spontaneous morphine withdrawal

The expression of specific genes is an important factor of neuroplastic changes during the formation of opiate dependence. Neurotrophic factors participate in structural-functional modifications in CNS regions after opiate intoxication. Here, we studied the levels of mRNAs of the brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and insulin-like growth factor 1 (IGF1) in the frontal cortex, striatum, hippocampus, and midbrain after spontaneous morphine withdrawal in dependent rats. To induce physical dependence, morphine was injected intraperitoneally twice a day with increasing doses of 10–100 mg/kg for 6 days. The expression of mRNAs of BDNF, NGF, and IGF1 in the brain areas was estimated 40 hours after spontaneous morphine withdrawal using the real-time PCR method. We found that spontaneous morphine withdrawal induced an elevation of BDNF and IGF1 mRNAs in the frontal cortex. In the hippocampus and the midbrain only BDNF mRNA increased. The content of NGF mRNA did not change in all regions studied. We believe that changes in the expression of BDNF and IGF1 are involved in the mechanisms of neuroplastic modification during the formation of opiate dependence.     

Effect of the novel sigma1 receptor ligand and putative atypical antipsychotic E-5842 on BDNF mRNA expression in the rat brain

Changes in the expression of brain-derived neurotrophic factor (BDNF) have been implicated in some neuropsychiatric disorders. Several antipsychotic drugs affect the expression of BDNF mRNA in different areas of the rat brain. We examined the effect of single or repeated administration of 4-[4-fluorophenyl]-1,2,3,6-tetra-hydo-1-[4-[1,-2,4-triazol-1-il]butyl]pyridine citrate) (E-5842), a sigma1 receptor ligand and putative atypical antipsychotic drug on the expression of BDNF mRNA in rats. Acute treatment with E-5842 induced a down-regulation of BDNF mRNA levels in the frontal cortex and hippocampus, while a chronic treatment had no effect. Levels of another neurotrophin, nerve growth factor (NGF), remained unaltered after either acute or chronic treatment. The effects suggest that any therapeutic properties of E-5842 are not mediated by stimulation of BDNF or NGF, whereas the regulation of these trophic factors may be part of the mechanism of action of sigma1 receptor ligands.     

Function and evolution in the NGF family and its receptors.

The gene family of neurotrophins includes nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4). Recently, neurotrophin-5 (NT-5), a possible mammalian homologue to NT-4 described in the frog Xenopus, has been cloned in man and rat. The neurotrophins stimulate survival and differentiation of a range of target neurons by binding to cell surface receptors. The structure of NGF has recently been clarified from crystallographic data. The similarities between the different neurotrophins are substantial with the variable regions, giving specificity to each of the family members, being localized to some exposed loop regions. Low-affinity binding (Kd of 10(-9) M) of all tested neurotrophins is mediated via a 75 K glycoprotein (LNGFR) that has been cloned and characterized. A 140 K tyrosine protein kinase encoded by the proto-oncogene trk has been found to bind NGF with high affinity (Kd of 10(-11) M) and to evoke the cellular neurotrophic responses. In addition, a protein encoded by the trk-related gene trkB has been shown to bind BDNF. Recently, a third member of the trk family, trkC, has been cloned and demonstrated to function as a high-affinity receptor for NT-3. The expression of trk and LNGFR mRNA are co-localized in the rat brain to the medial septal nucleus and the nucleus of Broca's diagonal band containing the NGF-responsive magnocellular cholinergic neurons projecting to hippocampus and cerebral cortex. In sharp contrast, the pattern of expression of trkB is widely spread in many areas of the cortex as well as lateral septum. The trkB protein might serve general functions in large areas of the cortex. Site-directed mutagenesis and expression of recombinant chimaeric neurotrophin proteins have made it possible to localize a likely region for the interaction between NGF and the LNGFR. This region could be altered, resulting in the total loss of LNGFR binding by the mutant NGF protein without affecting the binding to the trk receptor which was sufficient for the full biological activity. Cladistic analysis of likely phylogenies within the neurotrophins shows BDNF and NT-4 to be most closely related whereas NGF may be the sister group to NT-3, BDNF, and NT-4. Neurotrophins offer obvious clinical possibilities for treatment of neurodegenerative diseases.     

Regulation of Neuropeptides in Adult Rat Forebrain by the Neurotrophins BDNF and NGF

The expression of neuropeptides and neurotrophic factors is altered in the hippocampus after seizure induction in rats. Because the increase in brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) mRNAs precede changes in neuropeptide expression after seizure, it is possible that BDNF and NGF mediate subsequent alterations in peptide expression. To test this hypothesis directly, BDNF or NGF was infused into the hippocampus and cortex of adult rats. To ascertain the regional specificity of any observed effects of neurotrophin administration on neuropeptide expression, infusions into the striatum were also studied. To control for specificity, vehicle was also infused into the same sites. Peptide and mRNA alterations were assessed by Northern analysis, immunohistochemistry and radioimmunoassay. BDNF produced elevations of peptide and mRNA for neuropeptide Y and cholecystokinin in hippocampus and cortex, and somatostatin in cortex. BDNF increased mRNAs for neuropeptide Y, cholecystokinin, substance P and dynorphin in striatum. In contrast, BDNF decreased dynorphin peptide and mRNA in hippocampus. NGF's effects were limited to small mRNA increases, without corresponding changes in peptide levels, for neuropeptide Y in hippocampus and striatum, substance P in cortex and cholecystokinin in striatum. The distinct and limited effects of NGF infusion on neuropeptide expression demonstrate that BDNF's effects are not non-specific results of protein infusion into the brain. These findings indicate that BDNF may play a regionally specific role in modulating neuropeptide expression in the normal brain as well as in various pathophysiological states.     

Repeated episodic exposure to ethanol affects neurotrophin content in the forebrain of the mature rat

Chronic exposure to ethanol can cause deficits in learning and memory. It has been suggested that withdrawal is potentially more damaging than the ethanol exposure per se. Therefore, we explored the effect of repeated episodic exposure to ethanol on key regulators of cortical activity, the neurotrophins. Rats were exposed to ethanol via a liquid diet for 3 days per week for 6-24 weeks. Control rats were pair-fed an isocaloric liquid diet or ad libitum fed chow and water. The concentrations of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) were determined using enzyme-linked immunosorbant assays (ELISAs). Five telencephalic structures were examined: parietal cortex, entorhinal cortex, hippocampus, the basal nucleus, and the septal nuclei. All five areas expressed each of the three neurotrophins; BDNF was most abundant and NGF the least. The parietal cortex was susceptible to ethanol exposure, NGF and BDNF content increased, and NT-3 content fell, whereas no changes were detectable in the entorhinal cortex. In the hippocampus, the amount all three neurotrophins increased following episodic ethanol exposure. Neurotrophin content in the two segments of the basal forebrain was affected; NGF and NT-3 content in the basal forebrain was reduced and NGF and BDNF content in the septal nuclei was increased by ethanol exposure. In many cases where ethanol had an effect, the change was transient so that by 24 weeks of episodic exposure, no significant changes were evident. Thus, the effects of ethanol are site- and time-dependent. This pattern differs from changes caused by chronic ethanol exposure, hence, neurotrophins must be vulnerable to the effects of withdrawal. Furthermore, the ethanol-induced changes do not appear to fit a model consistent with retrograde regulation, rather they suggest that neurotrophins act through autocrine/paracrine systems.     

Neurotrophin and neuropeptide expression in mouse brain is regulated by knockout of the norepinephrine transporter

Neurotrophins [e.g. nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3)] and neuropeptides such as corticotropin-releasing factor (CRF) are reported to contribute to the action of antidepressants (ADs). Norepinephrine transporter (NET) knockout (NETKO) mice represent a model of chronic AD treatment. In the present study, we examined brain-region-specific regulations of NT-3, NGF, BDNF and CRF at the mRNA and protein level in NET wild-type (NETWT) and NETKO mice by means of quantitative real-time PCR (qPCR) and two-site enzyme-linked immunosorbent assays (ELISAs), respectively. NETKO-induced changes were detected for NT-3 in olfactory bulb, brainstem and whole brain at the mRNA and for olfactory bulb at the protein level, for NGF mRNA and protein in olfactory bulb, cerebellum and brainstem and for CRF mRNA and protein in the hippocampus. In contrast, BDNF levels remained unaltered. Our results suggest that NETKO mice represent a useful model to examine gene regulation of downstream targets potentially involved in the action of ADs. We could delineate NT-3, NGF and CRF as being regulated in distinct brain regions by KO of the NET.     

Presence of brain-derived neurotrophic factor in brain and human and rat but not mouse serum detected by a sensitive and specific immunoassay

Brain-derived neurotrophic factor (BDNF) is one of several endogenous proteins that play key roles in neuronal development and homeostasis. We describe here the characterization and use of a sensitive and specific enzyme-linked immunoassay (EIA) for BDNF protein. Recombinant BDNF was detected at concentrations as low as 10 pg/ml, whereas the EIA did not detect NT-3, NT-4/5, or NGF at concentrations as high as 100 ng/ml. Because BDNF protein sequences are identical among humans, mice, and rats, we utilized the BDNF EIA to detect BDNF in the circulation or brain regions of these species. High concentrations of BDNF were detected in human and rat serum, and up to 50-fold lower BDNF levels were present in citrated human or rat plasma. The BDNF signal (66–141 pg/ml) in 20% human plasma was completely blocked by pre-exposure of plasma to a monoclonal antibody (Mab) specific for BDNF but not by exposure to 5-fold greater concentrations of an irrelevant Mab of the same isotype (IgGl). There was a significant and positive correlation (r = +0.86) between plasma levels of BDNF and serotonin, an indoleamine that is specifically released from activated platelets. These results are consistent with the view that the BDNF detected in human and rat plasma is derived from platelet degranulation, and that circulating levels of BDNF are negligible. In contrast to human or rat serum, mouse serum contained no detectable BDNF. However, BDNF protein was readily detectable at 108–256 ng/g of tissue in hippocampus, frontal cortex, and neostriatum of mice and rats. Thus, the failure to detect BDNF in murine serum was not due to an assay defect but highlights a significant species difference in the tissue-specific expression of BDNF that may be of biological importance. The presence of BDNF protein in blood and brain regions at quantities which greatly exceed those described for NGF confirm the abundant distribution of this broadly-acting neurotrophic factor.