Enhanced synthesis of brain-derived neurotrophic factor in the lesioned peripheral nerve: different mechanisms are responsible for the regulation of BDNF and NGF mRNA.

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are molecules which regulate the development and maintenance of specific functions in different populations of peripheral and central neurons, amongst them sensory neurons of neural crest and placode origin. Under physiological conditions NGF is synthesized by peripheral target tissues, whereas BDNF synthesis is highest in the CNS. This situation changes dramatically after lesion of peripheral nerves. As previously shown, there is a marked rapid increase in NGF mRNA in the nonneuronal cells of the damaged nerve. The prolonged elevation of NGF mRNA levels is related to the immigration of activated macrophages, interleukin-1 being the most essential mediator of this effect. Here we show that transsection of the rat sciatic nerve also leads to a very marked increase in BDNF mRNA, the final levels being even ten times higher than those of NGF mRNA. However, the time-course and spatial pattern of BDNF mRNA expression are distinctly different. There is a continuous slow increase of BDNF mRNA starting after day 3 post-lesion and reaching maximal levels 3-4 wk later. These distinct differences suggest different mechanisms of regulation of NGF and BDNF synthesis in non-neuronal cells of the nerve. This was substantiated by the demonstration of differential regulation of these mRNAs in organ culture of rat sciatic nerve and Schwann cell culture. Furthermore, using bioassays and specific antibodies we showed that cultured Schwann cells are a rich source of BDNF- and ciliary neurotrophic factor (CNTF)-like neurotrophic activity in addition to NGF. Antisera raised against a BDNF-peptide demonstrated BDNF-immunoreactivity in pure cultured Schwann cells, but not in fibroblasts derived from sciatic nerve.

Identification of cells in rat brain and peripheral tissues expressing mRNA for members of the nerve growth factor family.

Cells expressing mRNA for hippocampus-derived neurotrophic factor (HDNF/NT-3) or brain-derived neurotrophic factor (BDNF) were identified by in situ hybridization. In the rat brain, HDNF mRNA was predominantly found in pyramidal neurons in CA1 and CA2 of the hippocampus. Lower levels of HDNF mRNA were found in granular neurons of the dentate gyrus and in neurons of the taenia tecta and induseum griseum. BDNF mRNA-expressing cells were more widely distributed in the rat brain, with high levels in neurons of CA2, CA3, and the hilar region of the dentate gyrus, in the external and internal pyramidal layers of the cerebral cortex, in the claustrum, and in one brainstem structure. Lower levels were seen in CA1 and in the granular layer of the hippocampus, in the taenia tecta, and in the mammillary complex. In peripheral tissues, HDNF mRNA was found in glomerular cells in the kidney, secretory cells in the male rat submandibular gland, and epithelial cells in secondary and tertiary follicles in the ovary. Cells expressing BDNF mRNA were found in the dorsal root ganglia, where neurons of various sizes were labeled.

Loss of p53 but not ARF accelerates medulloblastoma in mice heterozygous for patched.

Brain malignancies represent the most common solid tumors in children, and they are responsible for significant mortality and morbidity. The molecular basis of the most common malignant pediatric brain tumor, medulloblastoma, is poorly understood. Mutations in several genes including the human homologue of the Drosophila segment polarity gene, patched (PTCH), the adenomatous polyposis coli gene (APC), beta-catenin, and p53 have been reported in subsets of hereditary and sporadic medulloblastoma. Inactivation of one Ptc allele in mice results in a 14% incidence of medulloblastoma. Here, we report a dramatic increase in the incidence (>95%) and accelerated development (prior to 12 weeks of age) of medulloblastoma in mice heterozygous for Ptc that lack p53. The acceleration of tumorigenesis in Ptc+/- mice is specific for loss of p53, because no change in tumor incidence was observed in Ptc+/- mice carrying a mutation in APC (Min+/-) or in Ptc+/- mice deficient in p19ARF. Thus, there is a specific interaction between p53 loss and heterozygosity of Ptc that results in medulloblastoma. This may be a consequence of increased genomic instability associated with loss of p53 function that may enhance the rate of acquisition of secondary mutations. Ptc+/- p53-/- mice provide a useful model for investigation of the molecular bases of medulloblastoma and for evaluation of the efficacy of therapeutic intervention strategies in a spontaneously arising endogenous brain tumor.

The normal patched allele is expressed in medulloblastomas from mice with heterozygous germ-line mutation of patched.

Defects in a developmental signaling pathway involving mammalian homologues of the Drosophila segment polarity gene, patched (ptc) and its ligand, sonic hedgehog (shh), contribute to tumor formation in several tissues. Recently, a subset of medulloblastoma, the most common malignant brain tumor in children, was found to contain somatic mutations in the human ptc gene. In addition, basal cell nevus syndrome (BCNS), or Gorlin syndrome, which is characterized by developmental anomalies and a predisposition to skin and nervous system malignancies, is associated with germ-line mutation of ptc. Targeted disruption of both alleles of ptc in mice results in embryonic lethality. However, ptc+/- mice survive and develop spontaneous cerebellar brain tumors, suggesting that ptc may function as a tumor suppressor gene. Therefore, we investigated ptc+/-mice as a model for human medulloblastoma. We report that 14% of ptc+/- mice develop central nervous system tumors in the posterior fossa by 10 months of age, with peak tumor incidence occurring between 16 and 24 weeks of age. The tumors exhibited several characteristics of human medulloblastoma, including expression of intermediate filament proteins specific for neurons and glia. Full-length ptc mRNA was present in all tumors analyzed, indicating that there was no loss of heterozygosity at the ptc locus. Nucleotide sequence of ptc mRNA from four tumors failed to identify any mutations. However, a comparison of the normal ptc sequence from C57BL/6 and 129Sv mice did reveal several polymorphisms. High levels of glil mRNA and protein were detected in the tumors, suggesting that the shh/ptc pathway was activated despite the persistence of ptc expression. These data indicate that haploinsufficiency of ptc is sufficient to promote oncogenesis in the central nervous system.

Initial studies of embryonic transplants of human hippocampus and cerebral cortex derived from schizophrenic women.

Human fetal brain tissue was obtained from first-trimester elective abortions of two women who also had schizophrenia. Portions of the embryonic hippocampus or cerebral cortex were transplanted into the anterior eye chamber of immunologically compromised athymic nude rats. In this environment, embryonic brain tissue derived from normal women generally continues organotypic growth and development for many months. Although initial survival after transplantation was normal, the tissue derived from schizophrenic women manifested less robust growth. However, cells in the transplants showed typical neuronal differentiation, with development of different neuronal types, such as pyramidal cells, granule cells, and gamma-aminobutyric acid (GABA)-containing interneurons. Rhythmic electrical activity was also observed, indicative of some local synaptic organization. The presence of messenger RNA (mRNA) for brain-derived neuronotrophic factor (BDNF) was observed using in situ hybridization. The reason for the decreased rate of growth of these transplants remains unknown and the significance of the finding cannot be assessed from only two fetuses. However, these preliminary findings suggest that fetal transplants may be a useful model system for the detection of developmental pathogenic processes in the expression and transmission of schizophrenia.

Neuronal and nonneuronal expression of neurotrophins and their receptors in sensory and sympathetic ganglia suggest new intercellular trophic interactions.

Nerve growth factor promotes the survival of populations of sensory and sympathetic neurons. Although ganglia have been used for classical assays of neurotrophin action, knowledge is incomplete regarding the spatial arrangements through which neurotrophins are delivered to responsive cells within the ganglia and their attached nerve trunks. Whereas populations of ganglionic neurons may be capable of responding to a particular neurotrophin in vitro, the spectrum of receptor components and neurotrophins expressed by the various neuronal and nonneuronal cells comprising the ganglia in adult rats remains to be elucidated in vivo. Brain-derived neurotrophic factor (BDNF) mRNA was expressed by a population of small to medium sized neurons in all sensory ganglia except in the mesencephalic nucleus of the trigeminal nerve. Interestingly, BDNF immunoreactivity was detected in a more widespread population of neurons of these ganglia, as well as in scattered satellite cells of both sensory and sympathetic ganglia. These nonneuronal cells also expressed mRNA encoding a truncated form of the BDNF receptor, trkBtrunc, and full-length transcripts of trkB appeared to be confined to neuronal populations. Several other components of neurotrophin receptors (low-affinity neurotrophin receptor, trk, and trkC) were prominently expressed by different populations of neuronal cells in sympathetic and sensory ganglia, but they were not detected in nonneuronal cells. Neither nerve growth factor nor neurotrophin-3 mRNAs were detected in these ganglia. Unexpectedly, BDNF and trkBtrunc expression was detected in oligodendrocytes myelinating the central processes of sensory neurons. Schwann cells did not express detectable quantities of either entity, thereby establishing a dramatic boundary delineated by neurotrophin/neurotrophin receptor expression that coincided with the interface between the oligodendroglia of the central nervous system (CNS) and Schwann cells of the peripheral nervous system (PNS). Localization of BDNF expression to an additional population of nonneuronal cells--satellite cells within sensory and sympathetic ganglia--suggest a more extensive role for neurotrophic factors than originally encompassed by the target-derived neurotrophic-factor-concept paradigm. These data support the hypothesis of a possible autocrine or paracrine trophic interaction between populations of neuronal and nonneuronal cells in the peripheral nervous system. BDNF expression in oligodendrocytes but not in Schwann cells at the CNS/PNS junction may provide an additional means of maintaining cell-appropriate connections in the nervous system.

Detection and characterization of a sensory microganglion associated with the spinal accessory nerve: a scanning laser confocal microscopic study of the neurons and their processes.

The spinal accessory nerve has been generally thought to be a cranial nerve with purely motor function, innervating the trapezius and sternocleidomastoid muscles. The present study identified clusters of sensory neurons consistently associated with this cranial nerve in adult rats. Either a single microganglion or several dispersed microganglia were found that adhered to the spinal root of the nerve, to small vessels, or were free within the subarachnoid space. The neurons of the ganglion had axons that joined the spinal root of the nerve proximal to its exit from the skull. Additional branches appeared to have an intracranial distribution within the arachnoid of the brainstem and along its vessels. Several findings suggest that the function of the ganglion is sensory and not autonomic. First, the architectural features of neurons within the ganglion (including their size, pseudounipolar morphology, and the lack of synaptic contacts) are similar to those of neurons in other sensory ganglia. Second, substance P and calcitonin gene-related peptide coexist within neurons of the microganglion, whereas markers for the major transmitters found in autonomic ganglia in rats are absent. Third, the expression of peptides in neurons of the ganglion was sensitive to neonatal capsaicin treatment. Finally, neurons within the ganglion were filled with a retrogradely transported dye after injection of the dye into the cervical spinal cord. Although the function of the ganglion is not known, its features are consistent with a role in nociception from the muscles of the spinal accessory complex, and it may be involved in headaches that have an occipital distribution.

Regulation of brain-derived neurotrophic factor messenger RNA and protein at the cellular level in pentylenetetrazol-induced epileptic seizures.

We have examined the effects of pentylenetetrazol-induced epileptic seizures on brain-derived neurotrophic factor messenger RNA and protein and on the messenger RNA of its receptor in the rat. Pentylenetrazol, which acts at the picrotoxin recognition site of the GABAA receptor, was injected intraperitoneally and induced seizures by decreasing the inhibitory GABAergic activity. The effects of a single acute convulsive dose (50 mg/kg) of pentylenetetrazol were analysed at different time points by in situ hybridization or immunohistochemistry. Kindling was induced by daily subconvulsive injections (30 mg/kg) of pentylenetetrazol. At different time points during the kindling process, the messenger RNAs of brain-derived neurotrophic factor and trkB and the protein levels of brain-derived neurotrophic factor were analysed. We showed that brain-derived neurotrophic factor messenger RNA dramatically increased in neurons of the granule cell layer, piriform cortex and amygdala 3 h but not 6 h after an acute high dose of pentylenetetrazol, while brain-derived neurotrophic factor-like immunoreactivity was decreased in the granule cell layer and neurons of the hilus. The trkB messenger RNA was similarly increased 3 h and 6 h after the injection and returned to control levels after 24 h. The first change during the kindling development was seen after the first severe seizure: brain-derived neurotrophic factor messenger RNA was markedly increased in the piriform cortex and amygdala but not in the hippocampus. In fully kindled rats, which had several severe seizures, brain-derived neurotrophic factor messenger RNA and trkB messenger RNA were unaffected 3 h and 24 h after the last pentylenetetrazol injection. However, brain-derived neurotrophic factor-like immunoreactivity was markedly increased in the hippocampal formation 3 h, 24 h and three days after the last pentylenetetrazol injection, and still increased after 10 days. These results suggest that brain-derived neurotrophic factor may be involved in protection mechanisms after damage during seizures and in sprouting responses. The piriform cortex/amygdala seems to be an area of origin for the kindling development.

Comparative study of brain-derived neurotrophic factor messenger RNA and protein at the cellular level suggests multiple roles in hippocampus, striatum and cortex.

Brain-derived neurotrophic factor (BDNF) is important for the development and trophic support of several neuronal groups in the rat. In the present study, the distribution of BDNF messenger RNA was studied by in situ hybridization, and the cellular localization of BDNF protein was investigated with anti-peptide antibodies. Anatomical investigations were also made in animals with prolonged epileptic seizures which show an enhanced expression of BDNF messenger RNA. Major forebrain areas studied were the hippocampus, striatum and cortex. The messenger RNA coding for the putative high-affinity receptor, tyrosine kinase B, was also visualized using in situ hybridization with a probe specific for the full-length form. In the hippocampus, granule cells and pyramidal neurons expressed BDNF messenger RNA and BDNF-like immunoreactivity. Interneurons in dendritic layers did not show labelling with either method. Tyrosine kinase B messenger RNA was found within neurons in all these regions. In the medial septum-diagonal band, nucleus basalis and lateral hypothalamus, neurons with punctate cytoplasmic immunofluorescence were found, and neurons in the lateral septum were diffusely positive for BDNF. In striatum, positive labelling of medium-sized neurons was found with the antibody, whereas BDNF messenger RNA was only detectable during seizures. A laminar pattern of neuronal labelling for BDNF messenger RNA and protein was found in the neocortex. The analysis of the anatomical distribution of BDNF-producing cells suggests a number of possible cellular interactions. In the hippocampus, BDNF might act in an autocrine or paracrine manner for granule cells and pyramidal neurons, and, in addition, may serve as a signal from these principal cells to interneurons. BDNF could be a target-derived and a locally produced trophic factor for cholinergic neurons in the medial septum. The expression of BDNF in the striatum suggests that this factor could be a target-derived factor for dopaminergic neurons of substantia nigra and/or work as an autocrine/ paracrine factor within the striatum itself.

Brain-derived neurotrophic factor (BDNF) peptide antibodies: characterization using a Vaccinia virus expression system.

We describe and characterize a series of polyclonal antibodies, generated against amino acid sequences unique to various regions within pro- and mature brain-derived neurotrophic factor (BDNF), a member of the highly conserved nerve growth factor (NGF) family of neurotrophins. Synthetic peptides were coupled to carrier proteins in the presence of glutaraldehyde to restrict the host animals' immune response to epitopes that are compatible with aldehyde fixation. Initial screenings of the reactivity of the antisera were made on brain sections processed for immunohistochemistry after peptide injections into brain parenchyma. As a means of further characterizing these peptide antisera, we have evaluated the reactivity and specificity of the peptide antibodies in BHK cells expressing recombinant pro- and mature BDNF protein from a T7 RNA polymerase-driven Vaccinia virus system. Several of the antibodies strongly stained components of cells transfected with the BDNF gene but did not label wild-type cells nor cells containing only the expression vector. It has also been possible to detect differential compartmentalization of the BDNF protein at various stages of processing in the BHK cells, as well as in situ in cryostat sections of brain tissue, with antisera to the pro- and mature protein. We conclude that several of our antisera recognize not only the specific peptide immunogens but also what appears to be the corresponding protein native to neurons.

High and low affinity NGF receptor mRNAs in human foetal spinal cord and ganglia.

The cellular localization of mRNAs encoding the low affinity NGF receptor (here referred to as LANR) and the putative high affinity receptor for NGF, trk, have been studied in the human foetal spinal and sympathetic ganglia, and spinal cord, using in situ hybridization. The receptor mRNAs were highly expressed in the spinal and sympathetic ganglia, with most but not all neurons expressing both LANR and trk mRNA. Spinal nerve rootlets distal to the spinal ganglia expressed LANR but not trk mRNA, confirming the presence of the low affinity receptor in developing Schwann cells. In the spinal cord, LANR mRNA was found throughout the medial and lateral motor columns while, trk mRNA was detected in scattered cells in the dorsal aspect of developing grey matter.

Neurotrophin-3 and neurotrophin receptor immunoreactivity in peptidergic enteric neurons.

In the rat small intestine, neurotrophin-3 immunoreactivity was identified in ganglion cells and in processes mostly innervating the mucosa and occasionally the muscle layer and vasculature. The vast majority of neurotrophin-3 immunoreactive neurons contained vasoactive intestinal polypeptide (VIP), but not substance P or related tachykinin (SP/TK). Neurotrophin receptors visualized by pan-trk immunoreactivity were found in numerous ganglion cells of both plexuses and in nerve processes in the intestinal wall. Pan-trk submucosal neurons contained VIP (36%) or SP/TK-IR (47%). Pan-trk myenteric neurons contained VIP-IR (57%) or SP/TK (27%). Our data suggest that neurotrophin-3 and neurotrophin receptors may be involved in the maintenance of enteric neuronal circuits, transmission and phenotypic expression.

The nerve growth factor receptor gene is expressed in both neuronal and non-neuronal tissues in the human fetus.

In situ hybridization was used to study expression of beta-nerve growth factor receptor (NGF-R) mRNA in the early human fetus. In 8- to 12-week old fetuses, high labelling was found over motoneurons along the entire length of the lateral motor column. High levels of NGF-R mRNA were also seen over most developing nerve cell bodies in both the dorsomedial and ventrolateral part of the dorsal root ganglia. Lower, but clearly specific labelling was detected over a subpopulation of cells in Auerbach's plexus in the intestines. Evidence for a non-neuronal expression of NGF-R mRNA came from labelling over a subpopulation of cells in glomeruli of the kidney in a 12-week old human embryo. Myoblasts in skeletal muscle anlagen were labelled as well as cells along peripheral nerve. The widespread expression of NGF-R mRNA in the human fetus suggests that the NGF-R is important for development of a variety of different tissues of both neuronal and non-neuronal origin.

Trimethyltin exposure in the rat induces delayed changes in brain-derived neurotrophic factor, fos and heat shock protein 70.

Trimethyltin chloride (TMT) treatment in adult rats leads to limbic brain lesions that are detectable with classical neuropathological techniques 3 days after exposure. In particular, the hippocampal cells of the CA3c region are affected. The temporal and regional characteristics of TMT toxicity as reflected in changes of activity-dependent factors were studied in adult male Sprague-Dawley rats using quantitative in situ hybridization and immunohistochemistry. No significant alterations in the BDNF mRNA were detected in hippocampus and cerebral cortex 1 and 4 h after 8 mg TMT/kg. Three days after TMT, a significant increase in BDNF mRNA was detected in CA1, and increases in BDNF mRNA were also seen in cortical layers. An increase in BDNF hybridization signal was seen over scattered neurons within and outside CA3c at 3 days. Four h after 8 mg TMT/kg, BDNF immunoreactivity was reduced in the pyramidal cells of the CA3c and CA1 regions as well as in the dentate gyrus. No significant change in BDNF immunoreactivity was seen in hippocampus or cerebral cortex 3 days after TMT. BDNF interacts with the high-affinity receptor tyrosine kinase B (trkB). No immediate alteration in trkB mRNA was seen in hippocampus or cerebral cortex after 8 mg TMT/kg, while at 3 days trkB mRNA was significantly reduced in the CA3c pyramidal cell layer. No changes could be detected in neurotrophin-3 mRNA at either 1, 4 h or 3 days after TMT. Three days after 8 mg TMT/kg, a major induction of hsp70 mRNA occurred in a subset of neurons in the CA3c region, concomitant with an increased expression of c-fos mRNA as well as Fos protein in the hilar region of hippocampus. Hence, an early and transient decrease in BDNF appears to occur after TMT exposure, which is succeeded at 3 days by increases in BDNF, c-fos and hsp 70 mRNAs, concomitant with a decrease in trkB mRNA in regions known to be vulnerable to TMT. These results demonstrate that TMT causes a delayed, spatially restricted increase in activity-dependent gene expression, making TMT-induced disturbances an interesting model of neurodegenerative events.

16-year trends in elements of lichens at Theodore Roosevelt National Park, North Dakota.

An epiphytic lichen and a soil lichen in two very closely related genera (Parmelia sulcata and Xanthoparmelia chlorochroa, respectively) were sampled 16 years apart at Theodore Roosevelt National Park in North Dakota and measured for their elemental content. Mercury and cadmium decreased approximately 30% over the time period in both species. Sulfur decreased 8% in the epiphytic species, but increased 20% in the soil lichen. Factor analysis revealed that soil elements were higher in the soil lichen, indicating there was some soil contamination in that species. A relationship between iron and titanium was found only in the soil lichen. Sulfur and mercury were highly enriched in both species relative to the soil, which suggests that the atmosphere is a contributing source of these elements. New baseline values were calculated, 22 elements for both species, although it is not recommended that the soil lichen be sampled in the future.

Association between caffeine intake and bone mass among young women: potential effect modification by depot medroxyprogesterone acetate use.

UNLABELLED: This study assessed associations between habitual caffeine intake and bone mass among young women. Analyses of the entire study population revealed no significant associations, while analyses restricted to women using depot medroxyprogesterone acetate (DMPA) showed modest inverse associations between caffeine intake and bone mineral content (BMC). INTRODUCTION: Some previous investigations among postmenopausal women suggest an inverse relationship between caffeine intake and bone mass, yet studies of this association among young women are few. METHODS: The association between habitual caffeine intake and bone mass was evaluated prospectively in a population-based cohort of 625 females, aged 14 to 40 years, adjusting for relevant biological and lifestyle factors. Caffeinated beverage intake was self-reported, and bone mineral content (BMC) and bone mineral density (BMD) were measured at baseline and every 6 months throughout a 24-month follow-up period using dual-energy x-ray absorptiometry. RESULTS: Cross-sectional analyses revealed no significant differences in mean BMC or BMD at baseline. Mean percentage and absolute changes in BMC and BMD were not associated with caffeine use. Repeated measures analyses similarly showed no significant association between caffeine intake at baseline and mean BMC or BMD measured during follow-up. However, among women using depot medroxyprogesterone acetate (DMPA), modest inverse associations between caffeine and BMC (but not BMD) were detected. CONCLUSIONS: Our data suggest that heavy habitual consumption of caffeinated beverages does not adversely impact bone mass among young women in general. Greater caffeine intake may be associated with lower BMC among DMPA users.

Brain-derived neurotrophic factor and trkB receptor mRNAs in grafts of cortex cerebri.

Trophic factors are expressed by neurons throughout several areas of the CNS. We studied the mRNA expression of a member of the neurotrophin family, brain-derived neurotrophic factor (BDNF), and of the two receptor transcripts, full-length trkB and truncated trkB in single intraocular cortex cerebri grafts and in double cortex cerebri grafts. All single as well as double intraocular cortex grafts grew well, reaching a maximal size 4 weeks postgrafting. BDNF mRNA was moderately expressed in neurons in all intraocular grafts and significantly increased compared to that in adult rat cortex. Truncated trkB mRNA was strongly expressed in neurons and glia, while full-length trkB mRNA was moderately expressed only in neurons in the intraocular cortical grafts. The expression of the two trkB transcripts in the grafted cortex did not differ from that of adult rat cortex. No difference in the expression of mRNAs for full-length or truncated trkB was found between single grafts grown for 4 or 8 weeks or between single and double grafts. Similarly, no difference in expression of BDNF mRNA in single grafts grown for 4 or 8 weeks was detected. However, BDNF mRNA levels were significantly lower in grafts which were placed in close contact with previously grafted cortex in the eye chamber. Moreover, contact with a second graft led to downregulation of BDNF mRNA in the first graft.(ABSTRACT TRUNCATED AT 250 WORDS)

Cellular hybridization for BDNF, trkB, and NGF mRNAs and BDNF-immunoreactivity in rat forebrain after pilocarpine-induced status epilepticus.

The messenger RNAs (mRNAs) for the neurotrophins, brain-derived neurotrophic factor (BDNF), and nerve growth factor (NGF), are upregulated during epileptic seizure activity, as visualized by in situ hybridization techniques. Neurotrophins might be protective against excitotoxic cell stress, and the upregulation during seizures might provide such cell protection. In this study, a high dose of pilocarpine (300 mg/kg) was used to induce long-lasting, limbic motor status epilepticus and a selective pattern of brain damage. The regulation of BDNF, trkB, and NGF mRNA was studied by in situ hybridization at 1, 3, 6, and 24 h after induction of limbic motor status epilepticus. BDNF immunoreactivity was examined with an anti-peptide antibody and the neuropathological process studied in parallel. BDNF mRNA increased in hippocampus, neocortex, piriform cortex, striatum, and thalamus with a maximum at 3-6 h. Hybridization levels increased earlier in the resistant granule and CA1 cells as compared to the vulnerable CA3 neurons. BDNF immunoreactivity was elevated in dentate gyrus at 3-6 h. trkB mRNA increased in the entire hippocampus. NGF mRNA in hippocampus appeared in dentate gyrus at 3-6 h and declined in hilar neurons at 6-24 h. Cell damage was found in the CA3 area, entire basal cortex, and layers II/III of neocortex. Endogenous neurotrophins are upregulated during status epilepticus caused by pilocarpine, which is related to the coupling between neuronal excitation and trophic factor expression. This upregulation of neurotrophic factors may serve endogenous protective effects; however, the excessive levels of neuronal hyperexcitation resulting from pilocarpine seizures lead to cell damage which cannot be prevented by endogenous neurotrophins.

Localization of brain-derived neurotrophic factor mRNA to neurons in the brain by in situ hybridization.

Brain-derived neurotrophic factor (BDNF) which supports survival of and fiber outgrowth from sensory neurons and retinal ganglion cells has recently been cloned. It is closely related to nerve growth factor (NGF), which we had demonstrated earlier by in situ hybridization to be expressed in the brain by cholinergic target neurons. Using oligonucleotide probes we now describe aspects of the distribution of BDNF mRNA in the adult pig and rat brain. Similar to NGF, BDNF expression in vivo in the central nervous system appeared to be mostly confined to neurons. In the hippocampal formation, strongly labeled neurons were found in and around the pyramidal layer, as well as in the granular layer and the hilus of the dentate gyrus of both pig and rat. Other neurons in these areas were more weakly labeled or unlabeled. High BDNF expression was also shown in scattered neurons in cortex cerebri, in many neurons in claustrum and in certain other areas. These results demonstrate that the similarity in structure between NGF and BDNF is paralleled by a similarity in cellular expression, suggesting that BDNF, like NGF, may act via a direct neuron-to-neuron interaction. It is possible that some neurons, for example, in the hippocampal formation, express both NGF and BDNF. Finally, the localization of BDNF expression in cortical areas and in claustrum suggests that the target neurons for this factor extend beyond the sensory ganglion and retinal ganglion cells.