Semaphorin 3A growth cone collapse requires a sequence homologous to tarantula hanatoxin

Axonal guidance is key to the formation of neuronal circuitry. Semaphorin 3A (Sema 3A; previously known as semaphorin III, semaphorin D, and collapsin-1), a secreted subtype of the semaphorin family, is an important axonal guidance molecule in vitro and in vivo. The molecular mechanisms of the repellent activity of semaphorins are, however, poorly understood. We have now found that the secreted semaphorins contain a short sequence of high homology to hanatoxin, a tarantula K+ and Ca2+ ion channel blocker. Point mutations in the hanatoxin-like sequence of Sema 3A reduce its capacity to repel embryonic dorsal root ganglion axons. Sema 3A growth cone collapse activity is inhibited by hanatoxin, general Ca2+ channel blockers, a reduction in extracellular or intracellular Ca2+, and a calmodulin inhibitor, but not by K+ channel blockers. Our data support an important role for Ca2+ in mediating the Sema 3A response and suggest that Sema 3A may produce its effects by causing the opening of Ca2+ channels.

Genetically targeted cell disruption in Caenorhabditis elegans

The elimination of identified cells is a powerful tool for investigating development and system function. Here we report on genetically mediated cell disruption effected by the toxic Caenorhabditis elegans mec-4(d) allele. We found that ectopic expression of mec-4(d) in the nematode causes dysfunction of a wide range of nerve, muscle, and hypodermal cells. mec-4(d)-mediated toxicity is dependent on the activity of a second gene, mec-6, rendering cell disruption conditionally dependent on genetic background. We describe a set of mec-4(d) vectors that facilitate construction of cell-specific disruption reagents and note that genetic cell disruption can be used for functional analyses of specific neurons or neuronal classes, for confirmation of neuronal circuitry, for generation of nematode populations lacking defined classes of functional cells, and for genetic screens. We suggest that mec-4(d) and/or related genes may be effective general tools for cell inactivation that could be used toward similar purposes in higher organisms.

Transforming neural computations and representing time

Motifs of neural circuitry seem surprisingly conserved over different areas of neocortex or of paleocortex, while performing quite different sensory processing tasks. This apparent paradox may be resolved by the fact that seemingly different problems in sensory information processing are related by transformations (changes of variables) that convert one problem into another. The same basic algorithm that is appropriate to the recognition of a known odor quality, independent of the strength of the odor, can be used to recognize a vocalization (e.g., a spoken syllable), independent of whether it is spoken quickly or slowly. To convert one problem into the other, a new representation of time sequences is needed. The time that has elapsed since a recent event must be represented in neural activity. The electrophysiological hallmarks of cells that are involved in generating such a representation of time are discussed. The anatomical relationships between olfactory and auditory pathways suggest relevant experiments. The neurophysiological mechanism for the psychophysical logarithmic encoding of time duration would be of direct use for interconverting olfactory and auditory processing problems. Such reuse of old algorithms in new settings and representations is related to the way that evolution develops new biochemistry.

Gene-experience interaction alters the cholinergic septohippocampal pathway of mice

Spatial learning requires the septohippocampal pathway. The interaction of learning experience with gene products to modulate the function of a pathway may underlie use-dependent plasticity. The regulated release of nerve growth factor (NGF) from hippocampal cultures and hippocampus, as well as its actions on cholinergic septal neurons, suggest it as a candidate protein to interact with a learning experience. A method was used to evaluate NGF gene-experience interaction on the septohippocampal neural circuitry in mice. The method permits brain region-specific expression of a new gene by using a two-component approach: a virus vector directing expression of cre recombinase; and transgenic mice carrying genomic recombination substrates rendered transcriptionally inactive by a “floxed” stop cassette. Cre recombinase vector delivery into transgenic mouse hippocampus resulted in recombination in 30% of infected cells and the expression of a new gene in those cells. To examine the interaction of the NGF gene and experience, adult mice carrying a NGF transgene with a floxed stop cassette (NGFXAT) received a cre recombinase vector to produce localized unilateral hippocampal NGF gene expression, so-called “activated” mice. Activated and control nonactivated NGFXAT mice were subjected to different experiences: repeated spatial learning, repeated rote performance, or standard vivarium housing. Latency, the time to complete the learning task, declined in the repeated spatial learning groups. The measurement of interaction between NGF gene expression and experience on the septohippocampal circuitry was assessed by counting retrogradely labeled basal forebrain cholinergic neurons projecting to the hippocampal site of NGF gene activation. Comparison of all NGF activated groups revealed a graded effect of experience on the septohippocampal pathway, with the largest change occurring in activated mice provided with repeated learning experience. These data demonstrate that plasticity of the adult spatial learning circuitry can be robustly modulated by experience-dependent interactions with a specific hippocampal gene product.

A TSG101/MDM2 regulatory loop modulates MDM2 degradation and MDM2/p53 feedback control

The p53 tumor suppressor protein and the MDM2 oncoprotein form a feedback-control loop that up-regulates cellular MDM2 production, blocks p53 activity, and promotes p53 decay. tsg101 was discovered as a gene whose deficiency results in neoplastic transformation of NIH 3T3 cells and the ability to generate metastatic tumors in nude mice. Its protein product contains a domain, Ubc, characteristic of the catalytic domain of ubiquitin conjugase (E2) enzymes but lacking an active-site cysteine crucial for ubiquitin conjugase activity. Here we report that TSG101 participates with MDM2 in an autoregulatory loop that modulates the cellular levels of both proteins, and also of p53, by affecting protein decay. We show that the Ubc domain of TSG101 interferes with ubiquitination of MDM2, that TSG101 inhibits MDM2 decay and elevates its steady-state level, and that these events are associated with down-regulation of p53 protein. Conversely, pulse–chase and Western blot experiments in wild-type and mutant fibroblasts indicate that elevation of MDM2 by overexpression of wild-type p53, by amplification of the endogenous MDM2 gene, or by transfection of MDM2-expressing constructs promotes TSG101 loss, which we show occurs by 26S proteasome-dependent decay. Our results identify TSG101 as both a regulator of, and target of, MDM2/p53 circuitry.

Urocortin II: A member of the corticotropin-releasing factor (CRF) neuropeptide family that is selectively bound by type 2 CRF receptors

Here we describe the cloning and initial characterization of a previously unidentified CRF-related neuropeptide, urocortin II (Ucn II). Searches of the public human genome database identified a region with significant sequence homology to the CRF neuropeptide family. By using homologous primers deduced from the human sequence, a mouse cDNA was isolated from whole brain poly(A)+ RNA that encodes a predicted 38-aa peptide, structurally related to the other known mammalian family members, CRF and Ucn. Ucn II binds selectively to the type 2 CRF receptor (CRF-R2), with no appreciable activity on CRF-R1. Transcripts encoding Ucn II are expressed in discrete regions of the rodent central nervous system, including stress-related cell groups in the hypothalamus (paraventricular and arcuate nuclei) and brainstem (locus coeruleus). Central administration of 1–10 μg of peptide elicits activational responses (Fos induction) preferentially within a core circuitry subserving autonomic and neuroendocrine regulation, but whose overall pattern does not broadly mimic the CRF-R2 distribution. Behaviorally, central Ucn II attenuates nighttime feeding, with a time course distinct from that seen in response to CRF. In contrast to CRF, however, central Ucn II failed to increase gross motor activity. These findings identify Ucn II as a new member of the CRF family of neuropeptides, which is expressed centrally and binds selectively to CRF-R2. Initial functional studies are consistent with Ucn II involvement in central autonomic and appetitive control, but not in generalized behavioral activation.

Regional and cellular fractionation of working memory

This chapter recounts efforts to dissect the cellular and circuit basis of a memory system in the primate cortex with the goal of extending the insights gained from the study of normal brain organization in animal models to an understanding of human cognition and related memory disorders. Primates and humans have developed an extraordinary capacity to process information “on line,” a capacity that is widely considered to underlay comprehension, thinking, and so-called executive functions. Understanding the interactions between the major cellular constituents of cortical circuits—pyramidal and nonpyramidal cells—is considered a necessary step in unraveling the cellular mechanisms subserving working memory mechanisms and, ultimately, cognitive processes. Evidence from a variety of sources is accumulating to indicate that dopamine has a major role in regulating the excitability of the cortical circuitry upon which the working memory function of prefrontal cortex depends. Here, I describe several direct and indirect intercellular mechanisms for modulating working memory function in prefrontal cortex based on the localization of dopamine receptors on the distal dendrites and spines of pyramidal cells and on interneurons in the prefrontal cortex. Interactions between monoamines and a compromised cortical circuitry may hold the key to understanding the variety of memory disorders associated with aging and disease.

Anatomy of word and sentence meaning

Reading and listening involve complex psychological processes that recruit many brain areas. The anatomy of processing English words has been studied by a variety of imaging methods. Although there is widespread agreement on the general anatomical areas involved in comprehending words, there are still disputes about the computations that go on in these areas. Examination of the time relations (circuitry) among these anatomical areas can aid in understanding their computations. In this paper, we concentrate on tasks that involve obtaining the meaning of a word in isolation or in relation to a sentence. Our current data support a finding in the literature that frontal semantic areas are active well before posterior areas. We use the subject’s attention to amplify relevant brain areas involved either in semantic classification or in judging the relation of the word to a sentence to test the hypothesis that frontal areas are concerned with lexical semantics and posterior areas are more involved in comprehension of propositions that involve several words.

Genome projects and gene pools: New germplasm for plant breeding?

Crop gene pools have adapted to and sustained the demands of agricultural systems for thousands of years. Yet, very little is known about their content, distribution, architecture, or circuitry. The presumably shallow elite gene pools often continue to yield genetic gains while the exotic pools remain mostly untapped, uncharacterized, and underutilized. The concept and content of a crop’s gene pools are being changed by advancements in plant science and technology. In the first generation of plant genomics, DNA markers have refined some perceptions of genetic variation by providing a glimpse of a primary source, DNA polymorphism. The markers have provided new and more powerful ways of assessing genetic relationships, diversity, and merit by infusing genetic information for the first time in many scenarios or in a more comprehensive manner for others. As a result, crop gene pools may be supplemented through more rapid and directed methods from a greater variety of sources. Previously limited by the barriers of sexual reproduction, the native gene pools will soon be complemented by another gene pool (transgenes) and perhaps by other native exotic gene pools through comparative analyses of plants’ biological repertoire. Plant genomics will be an important force of change for crop improvement. The plant science community and crop gene pools may be united and enriched as never before. Also, the genomes and gene pools, the products of evolution and crop domestication, will be reduced and subjected to the vagaries and potential divisiveness of intellectual property considerations. Let the gains begin.

Neural codes: Firing rates and beyond

Computational neuroscience has contributed significantly to our understanding of higher brain function by combining experimental neurobiology, psychophysics, modeling, and mathematical analysis. This article reviews recent advances in a key area: neural coding and information processing. It is shown that synapses are capable of supporting computations based on highly structured temporal codes. Such codes could provide a substrate for unambiguous representations of complex stimuli and be used to solve difficult cognitive tasks, such as the binding problem. Unsupervised learning rules could generate the circuitry required for precise temporal codes. Together, these results indicate that neural systems perform a rich repertoire of computations based on action potential timing.

Plasticity in the neural coding of auditory space in the mammalian brain

Sound localization relies on the neural processing of monaural and binaural spatial cues that arise from the way sounds interact with the head and external ears. Neurophysiological studies of animals raised with abnormal sensory inputs show that the map of auditory space in the superior colliculus is shaped during development by both auditory and visual experience. An example of this plasticity is provided by monaural occlusion during infancy, which leads to compensatory changes in auditory spatial tuning that tend to preserve the alignment between the neural representations of visual and auditory space. Adaptive changes also take place in sound localization behavior, as demonstrated by the fact that ferrets raised and tested with one ear plugged learn to localize as accurately as control animals. In both cases, these adjustments may involve greater use of monaural spectral cues provided by the other ear. Although plasticity in the auditory space map seems to be restricted to development, adult ferrets show some recovery of sound localization behavior after long-term monaural occlusion. The capacity for behavioral adaptation is, however, task dependent, because auditory spatial acuity and binaural unmasking (a measure of the spatial contribution to the “cocktail party effect”) are permanently impaired by chronically plugging one ear, both in infancy but especially in adulthood. Experience-induced plasticity allows the neural circuitry underlying sound localization to be customized to individual characteristics, such as the size and shape of the head and ears, and to compensate for natural conductive hearing losses, including those associated with middle ear disease in infancy.

GABAergic feedforward projections from the inferior colliculus to the medial geniculate body.

A novel and robust projection from gamma-aminobutyric acid-containing (GABAergic) inferior colliculus neurons to the media] geniculate body (MGB) was discovered in the cat using axoplasmic transport methods combined with immunocytochemistry. This input travels with the classical inferior colliculus projection to the MGB, and it is a direct ascending GABAergic pathway to the sensory thalamus that may be inhibitory. This bilateral projection constitutes 10-30% of the neurons in the auditory tectothalamic system. Studies by others have shown that comparable input to the corresponding thalamic visual or somesthetic nuclei is absent. This suggests that monosynaptic inhibition or disinhibition is a prominent feature in the MGB and that differences in neural circuitry distinguish it from its thalamic visual and somesthetic counterparts.

A presynaptic locus for long-term potentiation of elementary synaptic transmission at mossy fiber synapses in culture.

The complex circuitry of the CA3 region and the abundance of collateral connections has made it difficult to study the mossy fiber pathway in hippocampal slices and therefore to establish the site of expression of long-term potentiation at these synapses. Using a novel cell culture system, we have produced long-term potentiation of the elementary synaptic connections on single CA3 pyramidal neurons following tetanic stimulation of individual dentate gyrus granule cells. As is the case for the hippocampal slice, this potentiation was independent of N-methyl-D-aspartate receptor activation, was simulated by application of forskolin, and its induction did not require any modulatory input. The increase in synaptic strength was accompanied by a reduction in the number of failures of transmission and by an increase in the coefficient of variation of the responses and was prevented by presynaptic injection of an inhibitor of protein kinase A. These findings show that mossy fiber long-term potentiation has a presynaptic locus and that its expression is dependent on protein kinase A.

An anatomical substrate for experience-dependent plasticity of the rat barrel field cortex.

The objective of this study was to examine the influence of sensory experience on the synaptic circuitry of the cortex. For this purpose, the quantitative distribution of the overall and of the gamma-aminobutyric acid (GABA) population of synaptic contacts was investigated in each layer of the somatosensory barrel field cortex of rats which were sensory deprived from birth by continuously removing rows of whiskers. Whereas there were no statistically significant changes in the quantitative distribution of the overall synaptic population, the number and proportion of GABA-immunopositive synaptic contacts were profoundly altered in layer IV of the somatosensory cortex of sensory-deprived animals. These changes were attributable to a specific loss of as many as two-thirds of the GABA contacts targeting dendritic spines. Thus, synaptic contacts made by GABA terminals in cortical layer IV and, in particular, those targeting dendritic spines represent a structural substrate of experience-dependent plasticity. Furthermore, since in this model of cortical plasticity the neuronal receptive-field properties are known to be affected, we propose that the inhibitory control of dendritic spines is essential for the elaboration of these functional properties.

The site of action of neuronal acidic fibroblast growth factor is the organ of Corti of the rat cochlea.

Here we show that the mature cochlear neurons are a rich source of acidic fibroblast growth factor (aFGF), which is expressed in the neuronal circuitry consisting of afferent and efferent innervation. The site of action of neuronal aFGF is likely to reside in the organ of Corti, where one of the four known FGF receptor (FGFR) tyrosine kinases--namely, FGFR-3 mRNA--is expressed. Following acoustic overstimulation, known to cause damage to the organ of Corti, a rapid up-regulation of FGFR-3 is evident in this sensory epithelium, at both mRNA and protein levels. The present results provide in vivo evidence for aFGF being a sensory neuron-derived, anterogradely transported factor that may exert trophic effects on a peripheral target tissue. In this sensory system, aFGF, rather than being a neurotrophic factor, seems to promote maintenance of the integrity of the organ of Corti. In addition, aFGF, released from the traumatized nerve endings, may be one of the first signals initiating protective recovery and repair processes following damaging auditory stimuli.