Distribution and retrograde transport of trophic factors in the central nervous system: functional implications for the treatment of neurodegenerative diseases.

Neurotrophins play a crucial role in the maintenance, survival and selective vulnerability of various neuronal populations within the normal and diseased brain. Several families of growth promoting substances have been identified within the central nervous system (CNS) including the superfamily of nerve growth factor related neurotrophin factors, glial derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF). In addition, other non-neuronal growth factors such as fibroblast growth factor (FGF) have also been identified. This article reviews the trophic anatomy of these factors within the CNS. Intraventricular and intraparenchymal injections of exogenous nerve growth factor result in retrograde labeling mainly within the cholinergic basal forebrain. Distribution of brain derived neurotrophic factor (BDNF) following intraventricular injection is minimal due to the binding to the trkB receptor along the ventricular wall. In contrast, intraparenchymal injections of BDNF results in widespread retrograde transport throughout the CNS. BDNF has also been shown to be transported anterogradely within the CNS. Infusion of GDNF into the CNS results in retrograde transport limited to the nigrostriatal pathway. Hippocampal injections of NT-3 retrogradely label mainly basal forebrain neurons. Retrograde transport of radiolabeled CNTF has only been observed in sensory neurons of the sciatic nerve. Following intraventricular and intraparenchymal infusion of radiolabeled bFGF, retrograde neuronal labeling was found in the telecephalon, diencephalon, mesencephalon and pons. In contrast retrograde labeling for aFGF was found only in the hypothalamus and midbrain. Since select neurotrophins traffic anterogradely and retrogradely within the nervous system, these proteins could be used to treat neurological diseases such as Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis.

NADPH-diaphorase/nitric oxide synthase-positive elements in the human olfactory bulb.

The distribution and morphology of nitric oxide-synthesizing elements in the human olfactory bulb were studied using NADPH-diaphorase histochemistry and nitric oxide synthase immunohistochemistry. NADPH-diaphorase was detected in all olfactory fibers and groups of superficial short-axon cells, deep short-axon cells, stellate cells and abundant centrifugal fibers. Similar cell types were nitric oxide synthase immunoreactive but olfactory fibers were immunonegative. The distribution patterns of nitric oxide-synthesizing elements showed significant differences from what has been reported in the olfactory bulb of macrosmatic mammals including rodents and insectivores. These differences are likely to correlate with interspecies differences in the processing of olfactory information.

Tyrosine kinase A, galanin and nitric oxide synthase within basal forebrain neurons in the rat.

Cholinergic basal forebrain neurons appear to play a key role in cognition and attention. In rat, basal forebrain neurons express multiple proteins including the high-affinity signal transducing tyrosine kinase A receptor for nerve growth factor, the neuropeptide galanin and nitric oxide synthase, a marker for the novel neurotransmitter nitric oxide. The present study was undertaken to define the relationship between neurons expressing each of these markers within the medial septum-vertical limb of the diagonal band, horizontal limb of the diagonal band and nucleus basalis in colchicine pre-treated rats. Tyrosine kinase A-immunopositive neurons were seen throughout all subfields of the basal forebrain. In contrast, nitric oxide synthase- and galanin-immunoreactive neurons were mainly distributed within the septal-diagonal band complex. Co-localization experiments revealed that virtually all nitric oxide synthase-positive neurons (visualized by nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry) also contained tyrosine kinase A, whereas many fewer tyrosine kinase A neurons were nicotinamide adenine dinucleotide phosphate-diaphorase positive within the medial septum-vertical limb of the diagonal band. Within the horizontal limb of the diagonal band, numerous nicotinamide adenine dinucleotide phosphate-diaphorase neurons expressed tyrosine kinase A, whereas only a small number of tyrosine kinase A neurons contained nicotinamide adenine dinucleotide phosphate-diaphorase. Within the nucleus basalis very few neurons were nicotinamide adenine dinucleotide phosphate-diaphorase reactive, and a minor number contained tyrosine kinase A. Additional co-localization experiments revealed minor percentages of neurons containing nicotinamide adenine dinucleotide phosphate-diaphorase and galanin immunoreactivity within the various subfields of the basal forebrain. Within the horizontal limb of the diagonal band minor numbers of nicotinamide adenine dinucleotide phosphate-diaphorase-reactive perikarya displayed galanin. Similarly, only a few galanin-containing neurons expressed nicotinamide adenine dinucleotide phosphate-diaphorase. The existence of tyrosine kinase A, nitric oxide synthase and galanin within select neuronal subgroups of the cholinergic basal forebrain suggests that these perikarya are responsive to a complex set of chemical signals. A greater understanding of the chemical signature of the cholinergic basal forebrain neurons will provide the insight required to develop novel pharmacological approaches aimed at preventing or slowing the degenerative processes that effect these neurons in aging and pathologic disorders.

[The entorhinal cortex (hippocampal formation) in aging and Alzheimer's disease. Neuroanatomical interpretation]. / La corteza entorrinal (formación del hipocampo) en el envejecimiento y en la enfermedad de Alzheimer. Interpretación neuroanatómica.

This paper deals with the neuronal changes shown by the entorhinal cortex in aging (30 cases) and in Alzheimer's disease (17 cases). In both instances, changes show a neuronal loss (measured as an index of cortical atrophy). The entorhinal cortex more closely related to polymodal association cortex hardly shows any variation with age, while Alzheimer cases present very intense neuronal loss. This pattern is interpreted taking into account the present knowledge about the connectivity of the entorhinal cortex and the remainder of the hippocampal formation and their role in memory processing.

Decreased trkA gene expression within basal forebrain neurons in Alzheimer's disease.

In situ hybridization for TrkA mRNA was combined with quantitative optical densitometry to evaluate whether the expression of this gene is altered within cholinergic basal forebrain neurons (CBF) in Alzheimer's disease (AD). TrkA mRNA within individual nucleus basalis neurons was significantly reduced (66%) in AD cases relative to aged controls. Reverse transcription polymerase chain reaction quantitative analyses confirmed that TrkA mRNA levels decreased markedly in AD. In contrast, expression of the gene coding for for the low affinity p75NTR was not significantly altered in AD relative to aged controls. These data indicate that there is a selective defect in trkA gene expression in AD, supporting the hypothesis that the degeneration of CBF neurons seen in this disease results from impaired nerve growth factor trophic support.

Retrograde transport of brain-derived neurotrophic factor (BDNF) following infusion in neo- and limbic cortex in rat: relationship to BDNF mRNA expressing neurons.

Brain-derived neurotrophic factor (BDNF) was the second member of the nerve growth factor (NGF) family to be isolated. The ability of BDNF to be retrogradely transported following intraparenchymal infusion represents a unique neurobiological tool to determine the location of putative neuron-specific BDNF-responsive neuronal systems. In the present study, we infused recombinant human (rh) BDNF into the rodent neo- and limbic cortex and used a turkey anti-BDNF antibody to determine specific populations of neurons which retrogradely transport this neurotrophin. Frontal cortex infusion retrogradely labeled neurons within the ipsilateral and contralateral frontal cortex, basal forebrain, lateral hypothalamus, centrolateral, mediodorsal, ventrolateral, ventromedial, ventral posterior, rhomboid, reuniens, and medial geniculate thalamic nuclei, and locus coeruleus. Occipital cortex infusion retrogradely labeled neurons in the frontal, temporal, occipital, and perirhinal cortices as well as the claustrum, basal forebrain, thalamus, epithalamus, hypothalamus, and raphe nuclei. Dorsal hippocampal infusion retrogradely labeled neurons within the septal diagonal band, supramammillary nucleus, and entorhinal cortex and was also transported within various hippocampal subfields. Entorhinal cortex infusion retrogradely labeled neurons within the perirhinal cortex, endopiriform nucleus, piriform cortex, dentate gyrus, presubiculum, parasubiculum, CA1-CA4 fields, amygdaloid nuclei, basal forebrain, thalamus, hypothalamus, periaqueductal gray, raphe nuclei, and locus coeruleus. Amygdala infusion labeled neurons in the endopiriform nucleus, temporal cortex, piriform cortex, paralimbic cortex, hippocampus, subiculum, entorhinal cortex, amygdala, basal forebrain, thalamus, hypothalamus, substantia nigra, pars compacta, raphe, and pontine parabrachial nuclei. In situ hybridization experiments demonstrated that virtually all areas which retrogradely transport BDNF also express its message. Neuroanatomical distributional studies of BDNF will unravel specific central nervous system neurotrophic-responsive systems.

Intrastriatal and intraventricular infusion of brain-derived neurotrophic factor in the cynomologous monkey: distribution, retrograde transport and co-localization with substantia nigra dopamine-containing neurons.

The distribution and retrograde transport of brain-derived neurotrophic factor was examined using magnetic resonance imaging guided stereotaxic intracerebroventricular and intrastriatal infusion in the cynomologous monkey. Two intracerebroventricular animals were infused with brain-derived neurotrophic factor at a dose of 3 micrograms/h for 21 and 28 days. A third intracerebroventricular animal received sequential infusions of 15, 30 and 60 micrograms/h brain-derived neurotrophic factor each for seven days using an Alzet 2002 minipump. For the multiple intrastriatal animals (n = 5) a dose of 3 micrograms/h was infused into each site. One intrastriatal monkey was infused with vehicle solution of 10 mM phosphate-buffered saline pH 7.4 for 14 days resulting in no brain-derived neurotrophic factor immunoreactivity. Following the lower dose intracerebroventricular infusion, brain-derived neurotrophic factor immunoreactivity was confined to the ventricular ependymal layer. In the sequential higher dose intracerebroventricular case, the cannula was located mainly within the lateral ventricle, although there was damage to the ependymal wall and adjacent caudate nucleus. Brain-derived neurotrophic factor immunoreactivity revealed spread of injectate within the ipsilateral and to a lesser extent the contralateral caudate nucleus, septum, orbital cortex and ventricular ependymal wall. In this case, retrogradely labelled brain-derived neurotrophic factor neurons were found within the parafascicular thalamus and substantia nigra, pars compacta, as well as within cortex, vertical limb of the diagonal band and nucleus basalis. Brain-derived neurotrophic factor intrastriatal infusion retrogradely labelled perikarya within sensory motor cortex, parafascicular thelamus and substantia nigra, pars compacta. Sections from these cases dual-immunoreacted for brain-derived neurotrophic factor and tyrosine hydroxylase, the synthesizing enzyme for dopamine, revealed a subpopulation of pars compacta dopaminergic neurons which contained retrogradely transported brain-derived neurotrophic factor. These findings indicate that a select subgroup of nigral dopamine neurons retrogradely transport brain-derived neurotrophic factor in the primate. Furthermore it remains to be determined whether select nigral cells are responsive to the trophic influences of brain-derived neurotrophic factor in the normal and neuropathologic condition.

Reduced nicotinamide adenine dinucleotide phosphate-diaphorase/nitric oxide synthase profiles in the human hippocampal formation and perirhinal cortex.

Nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d)-stained profiles were evaluated throughout the human hippocampal formation (i.e., dentate gyrus, Ammon's horn, subicular complex, entorhinal cortex) and perirhinal cortex. NADPH-d staining revealed pleomorphic cells, fibers, and blood vessels. Within the entorhinal and the perirhinal cortices, darkly stained (type 1) NADPH-d pyramidal, fusiform, bipolar, and multipolar neurons with extensive dendrites were scattered mainly within deep layers and subjacent white matter. Moderately stained (type 2) NADPH-d round or oval neurons were seen mainly in layers II and III of the entorhinal and perirhinal cortices, in the dentate gyrus polymorphic layer, in the CA fields stratum pyramidal and radiatum, and in the subicular complex. The distribution of type 2 cells was more abundant in the perirhinal cortex compared to the hippocampal formation. Lightly stained (type 3) NADPH-d pyramidal and oval neurons were distributed in CA4, the entorhinal cortex medial subfields, and the amygdalohippocampal transition area. Sections concurrently stained for NADPH-d and nitric oxide synthase (NOS) revealed that all type 1 neurons coexpressed NOS, whereas types 2 and 3 were NOS immunonegative. NADPH-d fibers were heterogeneously distributed within the different regions examined and were frequently in close apposition to reactive blood vessels. The greatest concentration of fibers was in layers III and V-VI of the entorhinal and perirhinal cortices, dentate gyrus polymorphic and molecular layers, and CA1 and CA4. A band of fibers coursing within CA1 divided into dorsal and ventral bundles to reach the presubiculum and entorhinal cortex, respectively. Although the distribution of NADPH-d fibers was conserved across all ages examined (28-98 years), we observed an increase in the density of fiber staining in the aged cases. These results may be relevant to our understanding of selective vulnerability of neuronal systems within the human hippocampal formation in aging and in neurodegenerative diseases.

The human entorhinal cortex: a cytoarchitectonic analysis.

The entorhinal cortex of man is in the medial aspect of the temporal lobe. As in other mammalian species, it constitutes an essential component of the hippocampal formation and the route through which the neocortex interacts with the hippocampus. The importance of knowing its architecture in detail arises from the possibility of extrapolating it to experimental findings, notably in the nonhuman primate. We have investigated the cytoarchitectonic features of the human entorhinal cortex by using as a base our previous study (D.G. Amaral, R. Insausti, and W.M. Cowan [1987] J. Comp. Neurol. 264:326-355) of the nonhuman primate entorhinal cortex. We prepared serial sections of the temporal lobe from 35 normal brains. Thionin- and myelin-stained series were made of all cases. Sections spaced 500 microns apart through the full rostrocaudal extent of the entorhinal cortex were analyzed. The human entorhinal cortex is made up of six layers, of which layer IV does not appear throughout all subfields of the entorhinal cortex. The overall appearance resembles that of the adjacent neocortex in lateral and caudal portions. In harmony with general structural principles in the nonhuman primate entorhinal cortex, our analysis supports the partitioning of the human entorhinal cortex into eight different subfields. (1) The olfactory subfield (EO), the rostralmost field, is little laminated. (2) The lateral rostral subfield (ELr), laterally located, merges with the laterally adjacent perirhinal cortex. (3) The rostral subfield (ER) is between EO and ELr, with better differentiation of layers II and III than EO. (4) The medial intermediate subfield (EMI) is located at the medial border. (5) The intermediate field (EI) is a lateral continuation of EMI; lamina dissecans (layer IV) can be best appreciated in this field. (6) The lateral caudal subfield (ELc) laterally borders on EI as a continuation of ELr. (7) The caudal subfield (EC) lies caudal to the beginning of the hippocampal fissure, with a distinctive, clear space (Vc) between layers V and VI. (8) The caudal limiting field (ECL) forms the caudal termination of the entorhinal cortex. Thus our parcellation of the entorhinal cortex in man is largely parallel to that arrived at in the monkey. This close homology provides a rational basis for the application to clinical problems of anatomical and functional information obtained in experimental work in nonhuman primates.

TrkA-immunoreactive profiles in the central nervous system: colocalization with neurons containing p75 nerve growth factor receptor, choline acetyltransferase, and serotonin.

The present investigation used an antibody directed against the extracellular domain of the signal transducing nerve growth factor receptor, trkA, to reveal immunoreactive perikarya or fibers within the olfactory bulb and tubercle, cingulate cortex, nucleus accumbens, striatum, endopiriform nucleus, septal/diagonal band complex, nucleus basalis, hippocampal complex, thalamic paraventricular and reuniens nuclei, periventricular hypothalamus, interpeduncular nucleus, mesencephalic nucleus of the fifth nerve, dorsal nucleus of the lateral lemniscus, prepositus hypoglossal nucleus, ventral cochlear nucleus, ventral lateral tegmentum, medial vestibular nucleus, spinal trigeminal nucleus oralis, nucleus of the solitary tract, raphe nuclei, and spinal cord. Colocalization experiments revealed that virtually all striatal trkA-immunoreactive neurons (> 99%) coexpressed choline acetyltransferase (ChAT) but not p75 nerve growth factor receptor (NGFR). Within the septal/diagonal band complex virtually all trkA neurons (> 95%) coexpressed both ChAT and p75 NGFR. More caudally, dual stained sections revealed numerous trkA/ChAT (> 80%) and trkA/p75 NGFR (> 95%) immunoreactive neurons within the nucleus basalis. In the brainstem, raphe serotonergic neurons (45%) coexpressed trkA. Sections stained with a pan-trk antibody that recognizes primarily trkA, as well as trkB and trkC, labeled neurons within all of these regions as well as within the hypothalamic arcuate, supramammilary, and supraoptic nuclei, hippocampus, inferior and superior colliculus, substantia nigra, ventral tegmental area of T'sai, and cerebellular Purkinje cells. Virtually all of these other regions with the exception of the cerebellum also expressed pan-trk immunoreactivity in the monkey. The widespread expression of trkA throughout the central neural axis suggests that this receptor may play a role in signal transduction mechanisms linked to NGF-related substances in cholinergic basal forebrain and noncholinergic systems. These findings suggest that pharmacological use of ligands for trkA could have beneficial effects on the multiple neuronal systems that are affected in such disorders as Alzheimer's disease.

Intrastriatal infusions of brain-derived neurotrophic factor: retrograde transport and colocalization with dopamine containing substantia nigra neurons in rat.

The pattern of retrogradely transported BDNF, a member of the nerve growth family of neurotrophins, following intrastriatal infusion was immunohistochemically visualized within the rodent central nervous system. Human recombinant BDNF was infused at a rate of 3 micrograms/h for 7 days with an Alzet 2002 minipump prior to sacrifice. Tissue immunohistochemically processed using a turkey anti-BDNF antibody revealed retrogradely transported BNDF within neurons located mainly within the ipsilateral frontoparietal cortex (predominantly layer V), parafascicular and posterior thalamic nuclei, and substantia nigra, pars compacta. Sections dual immunoreacted for BNNF and tyrosine hydroxylase revealed a subpopulation of dopaminergic neurons (approximately 28%) within the pars compacta which contained retrogradely transported BDNF. Experiments in which a mixture of BDNF and the retrograde tracer fluorogold were simultaneously infused for 7 days into the striatum revealed BDNF and fluorogold single-labeled neurons as well as BDNF and fluorogold dual-labeled cells within the substantia nigra, pars compacta. These observations indicate that only a subpopulation of neurons within the substantia nigra retrogradely transport BDNF following intrastriatal infusion and thus only a subpopulation of cells may be responsive to the trophic influences of BDNF. The retrograde transport of trophins, such as BDNF, represents a unique neuroanatomical tool to selectivity map the location of specific neurotrophin-responsive systems. Unraveling the trophic anatomy of BDNF will aid in understanding its role in development, degeneration, and experimental animal models of regeneration providing essential data for its use in clinical neurodegenerative disorders including Parkinson's disease.

Cholinergic innervation in the human hippocampal formation including the entorhinal cortex.

The cholinergic innervation of the hippocampal formation is thought to play an important role in memory processes, but its organization in humans has not been described in detail. We studied the cholinergic innervation of the human hippocampal formation by means of immunohistochemistry with polyclonal antisera directed against acetylcholinesterase (AChE), choline acetyltransferase (ChAT), and the low-affinity (p75) nerve growth factor receptor (NGFR). The density of ChAT-like immunoreactive (ChAT-li) fibers differed substantially among the various regions, in general paralleling the pattern of AChE-li staining. One notable exception was the presence of AChE-li cell bodies. In contrast, ChAT immunoreactivity was associated only with fibers and terminals. NGFR-li staining corresponded closely to the ChAT-li fiber pattern. ChAT-li fibers in the CA fields diffusely filled the stratum pyramidale and extended into the stratum oriens and radiatum as well. The highest density was consistently observed in CA4 and CA3 subfields. Staining decreased from CA4 to CA1 and was substantially less dense in the subicular complex. In the entorhinal cortex, the ChAT- and NGFR-li fiber innervation displayed a laminar pattern, most intense over the nests of cells in layer II. There was a trend towards an age-related reduction in the density of ChAT- and AChE-li fibers and terminals. Nonetheless, we also found a surprisingly conserved NGFR-li innervation and the presence of occasional NGFR-li pyramidal cells, providing evidence of a plastic response in the brains of the elderly patients.

Parvalbumin and calbindin D-28K in the human entorhinal cortex. An immunohistochemical study.

Research is here reported on the distribution of immunoreactivities of the calcium-binding proteins parvalbumin and calbindin D-28K in the entorhinal cortex of normal human brains. Topographically, parvalbumin immunoreactive neurons were only seen in the lateral portion of the rostral entorhinal cortex, in continuity with the adjacent perirhinal cortex. The intermediate and caudal portions gave positive results along the mediolateral extension of the entorhinal cortex. The laminar distribution of parvalbumin immunoreactive neurons was similar throughout the entorhinal cortex. Heavy immunostaining, largely coincident with cell islands, was observed in cells and fibers in layer II, being densest in the deep half of layer III and more sparsely distributed in layers V and VI. Calbindin D-28K immunoreactivity was found throughout the entorhinal cortex. In contrast to parvalbumin immunoreactivity, calbindin D-28K was present from layer I up to upper layer III, the neurons being most numerous in the cell islands of layer II. These results show that rostromedial portions of the human entorhinal cortex contain calbindin immunoreactivity, but not parvalbumin, while the lateral, intermediate and caudal portions of the entorhinal cortex contain both calcium-binding proteins. As it is known that these two proteins belong to a subset of GABAergic neurons, we suggest that a topographical diversity in some of the cells may be responsible for inhibitory effects in the human entorhinal cortex. This proposed diversity might be relevant to the processing of information that the entorhinal cortex conveys to the dentate gyrus and receives from various components of the hippocampus, the subicular complex and other cortical and subcortical sources.