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This is a pilot study that examines how a person's gaze responds during the observation of a Japanese garden. Thirty-four participants were asked to viewed the same garden in three different ways: directly, through a pane of glass, or as a projected slide. We tracked their eye-movement during the first minutes of observation to observe different gaze strategies used in the different modalities of viewing. Our measurements show that when viewing the garden directly, participants focused on particular elements in their visual field. By contrast, when they viewed a slide they tended to scan the scene more and focus on all elements of the garden evenly. When viewing the real garden through a glass door, the responses were intermediate. Our results demonstrate that the visual evaluation of a space is different depending on the way in which the visual information is presented.

Myelination is a recent evolutionary addition that significantly enhances the speed of transmission in the neural network. Even slight defects in myelin integrity impair performance and enhance the risk of neurological disorders. Indeed, myelin degeneration is an early and well-recognized neuropathology that is age associated, but appears before cognitive decline. Myelin is only formed by fully differentiated oligodendrocytes, but the entire oligodendrocyte lineage are clear targets of the altered chemistry of the aging brain. As in neurons, unrepaired DNA damage accumulates in the postmitotic oligodendrocyte genome during normal aging, and indeed may be one of the upstream causes of cellular aging - a fact well illustrated by myelin co-morbidity in premature aging syndromes arising from deficits in DNA repair enzymes. The clinical and experimental evidence from Alzheimer's disease, progeroid syndromes, ataxia-telangiectasia and other conditions strongly suggest that oligodendrocytes may in fact be uniquely vulnerable to oxidative DNA damage. If this damage remains unrepaired, as is increasingly true in the aging brain, myelin gene transcription and oligodendrocyte differentiation is impaired. Delineating the relationships between early myelin loss and DNA damage in brain aging will offer an additional dimension outside the neurocentric view of neurodegenerative disease.

The development of oligodendrocytes, the myelinating cells of the vertebrate CNS, is regulated by a cohort of growth factors and transcription factors. Less is known about the signaling pathways that integrate extracellular signals with intracellular transcriptional regulators to control oligodendrocyte development. Cyclin dependent kinase 5 (Cdk5) and its co-activators play critical roles in the regulation of neuronal differentiation, cortical lamination, neuronal cell migration and axon outgrowth. Here we demonstrate a previously unrecognized function of Cdk5 in regulating oligodendrocyte maturation and myelination. During late embryonic development Cdk5 null animals displayed a reduction in the number of MBP+ cells in the spinal cord, but no difference in the number of OPCs. To determine whether the reduction of oligodendrocytes reflected a cell-intrinsic loss of Cdk5, it was selectively deleted from Olig1+ oligodendrocyte lineage cells. In Olig1Cre/+; Cdk5fl/fl conditional mutants, reduced levels of expression of MBP and PLP mRNA were observed throughout the CNS and ultrastructural analyses demonstrated a significant reduction in the proportion of myelinated axons in the optic nerve and spinal cord. Pharmacological inhibition or RNAi knockdown of Cdk5 in vitro resulted in the reduction in oligodendrocyte maturation, but had no effect on OPC cell proliferation. Conversely, over-expression of Cdk5 promoted oligodendrocyte maturation and enhanced process outgrowth. Consistent with this data, Cdk5−/− oligodendrocytes developed significantly fewer primary processes and branches than control cells. Together, these findings suggest that Cdk5 function as a signaling integrator to regulate oligodendrocyte maturation and myelination.

Initiation of a cell cycle in an adult neuron leads to cell death, placing great importance on the mechanisms that normally suppress the neuronal cell cycle. We have previously shown that the cyclin-dependent kinase Cdk5 is an important part of this process, but only when it is present in the nucleus. We report here that Cdk5 nuclear localization relies on its binding to the cyclin-dependent kinase inhibitor p27. Cdk5 has no intrinsic nuclear localization signal; in the absence of p27, two weak nuclear export signals that bind CRM1 cause it to shuttle to the cytoplasm. When a neuron is subjected to stress, such as exposure to β-amyloid, the Cdk5-p27 interaction is lost, reducing Cdk5 levels in the nucleus and depriving the neuron of a major cell cycle suppression mechanism. Caspase-3 is activated within hours, but death is not immediate; elevated levels of cytoplasmic Cdk5 appear to retard neuronal death by a mechanism that may involve Bcl2. These data suggest a model in which Cdk5 exerts a double protective function in neurons: chronically suppressing the cell cycle when located in the nucleus and transiently delaying cell death in the cytoplasm.

ATM is a PI3 kinase family member that plays an important role in DNA double strand break repair. ATM deficiency leads to ataxia-telangiectasia (AT), a syndrome involving cancer susceptibility, hypersensitivity to ionizing radiation, immune deficiency and sterility1, 2 – phenotypes that can straightforwardly be attributed to a defective response to DNA damage. Yet patients with AT also suffer from ataxia, speech defects and abnormal body movements3–5 – neurological phenotypes whose origins remain largely unexplained. Compounding the discordance, ATM mutations in mouse interfere with DNA repair but have only mild neurological symptoms6–9 suggesting that the link between DNA damage and the death of neurons and astrocytes can be broken10–12. We find that in neurons ATM protein has a significant cytoplasmic distribution. The functional importance of this cytoplasmic ATM is supported by our finding that in Atmtm1Awb mice long term potentiation in the Schaffer collateral pathway, induced by theta burst stimulation, is significantly reduced, as is the rate of vesicular dye release. ATM can be isolated from synaptosomes where it is found in a complex with two synaptic vesicle proteins: VAMP2 and synapsin-I, both of which must be phosphorylated to bind ATM. The neurological symptoms of ataxia-telangiectasia may thus result from defective cytoplasmic functions of ATM as well as defective DNA repair.

Cell cycle events have been documented to be associated with several human neurodegenerative diseases. This review focuses on two diseases – Alzheimer's disease and ataxia telangiectasia – as well as their mouse models. Cell cycle studies have shown that ectopic expression of cell cycle markers is spatially and regional correlated well with neuronal cell death in both disease conditions. Further evidence of ectopic cell cycling is found in both human diseases and in its mouse models. These findings suggest that loss of cell cycle control represents a common pathological root of disease, which underlies the defects in the affected brain tissues in both human and mouse. Loss of cell cycle control is a unifying hypothesis for inducing neuronal death in CNS. In the disease models we have examined, cell cycle markers appear before the more well-recognized pathological changes and thus could serve as early stress markers—outcome measures for preclinical trials of potential disease therapies. As a marker these events could serve as a new criterion in human pathological diagnosis. The evidence to date is compatible with the requirement for a second “hit” for a neuron to progress cell cycle initiation and DNA replication to death. If this were true, any intervention of blocking ‘second’ processes might prevent or slow the neuronal cell death in the process of disease. What is not known is whether, in an adult neuron, the cell cycle event is part of the pathology or rather a desperate attempt of a neuron under stress to protect itself.

The mouse Engrailed genes, En1 and En2, play an important role in the development of the cerebellum from its inception at the mid/hindbrain boundary in early embryonic development through cell type specification events and beyond. In the absence of En1, the cerebellum and caudal midbrain fail to develop normally--a phenotype that we have previously reported to be strain dependent. On the 129/S1 strain background, En1 null alleles lead to mid/hindbrain failure, whereas on the C57BL/6 background, En1 deficiency is compatible with near normal cerebellar development. We have pursued this dramatic effect of genetic background by performing a genetic modifier screen through F1 backcross and F1 intercross matings. The backcross has yielded two strong candidate intervals with suggestive linkage to a third region. Moreover, variations in rescue frequency among subgroups within the backcross indicate gender and parent of origin influences on rescue penetrance. The intercross data reveal locus heterogeneity of the En1 modifiers, with more than one compliment of C57BL/6 and 129/S1 alleles capable of mediating the rescue phenotype. These findings highlight the complexity and plasticity of gene networks involved in brain development.

We have sought to understand the regulation of the expression pattern of aldolase C (Zebrin II) in cerebellar Purkinje cells. Normally, aldolase C is expressed in a series of sagittal stripes of Purkinje cells interrupted by stripes of little or no expression. Genomic aldolase C:LacZ fusion genes with 1.8 kb of sequence 5' to the transcription start site drive CNS expression of LacZ only in astrocytes and cells of the pia mater. If the 5' portion of the transgene is extended to a full 5.0 kb, expression is reliably observed in Purkinje cells, yet none of the astrocyte expression is lost. We broke the additional 3.0 kb into 1.0 kb fragments and tested each for Purkinje cell enhancer activity when appended to the original 1.8 kb construct. We show that the 886 bp region from nucleotide -2796 to -3682 (relative to the start of transcription) contains virtually all of the Purkinje cell enhancer activity. However, neither the full 5.0 kb nor the 886 bp region directed a striped expression pattern, as is seen for the endogenous gene. Taken together, our study localizes a Purkinje cell enhancer to a small 5' region of the aldolase C gene and illustrates that the element(s) responsible for the normal anatomically complex pattern of aldolase C expression are separate from those conferring cell-type specificity. The relationship of these findings to previous work in other laboratories is discussed.

The vertebrate cerebellum is divided into a characteristic set of 13 parasagittal "bands" or modules that are revealed in many different domains-ranging from patterns of gene and protein expression to the organization of afferent input. We have used the expression of Zebrin II/aldolase C in Purkinje cells as a marker of these bands and have discovered several new features of their regulation. We find that appearance of the banded expression of aldolase C during development differs between rat and mouse. In agreement with previous reports there is, in rat, a transient period during which all Purkinje cells are positive for aldolase C expression. By contrast, in mouse, the pattern emerges in its adult (banded) form from the earliest postnatal times. This species difference is found in both mRNA and protein expression. There also appears to be a transition that occurs in vivo between postnatal days 8 and 10. Slice cultures established from cerebella at the younger age do not develop a complete banding pattern, even after 6 days in culture. Slice cultures established from postnatal day 10 mice develop the full pattern within 2 days. This difference cannot be overcome by manipulating the levels of neuronal activity in the cultures. Thus some event must occur in vivo that "releases" the adult pattern and allows it to be realized in the more artificial situation of the slice culture. Taken together the results offer a more complete picture of the regulation of the aldolase C gene in cerebellar Purkinje cells and suggest important species differences in its developmental expression pattern.

In the developing nervous system, neurons are generated at sites distant from their ultimate location. In the vertebrate CNS, neurons utilize distinct migration strategies to reach their ultimate residence. This review discusses the contribution of tangential migration to the architectural development of the cerebral cortex and cerebellum.

Mice completely deficient for Dvl1 , one of three mouse homologs of the Drosophila segment polarity gene Dishevelled , were created by gene targeting. Dvl1 - deficient mice are viable, fertile, and structurally normal. Surprisingly, these mice exhibited reduced social interaction, including differences in whisker trimming, deficits in nest-building, less huddling contact during home cage sleeping, and subordinate responses in a social dominance test. Sensorimotor gating was abnormal, as measured by deficits in prepulse inhibition of acoustic and tactile startle. Thus, Dvl1 mutants may provide a model for aspects of several human psychiatric disorders. These results are consistent with an interpretation that common genetic mechanisms underlie abnormal social behavior and sensorimotor gating deficits and implicate Dvl1 in processes underlying complex behaviors.

The cerebellum on the heterozygous (+/sg) staggerer mutant mouse has recently been proposed as a model system in which to study the genetic contribution to the normal process of central nervous system aging since there is significant loss of neurons from 3 to 12 months of age (Shojaeian-Zanjani, H., Mariani, J., Delhaye-Bouchaud, N., and Herrup, K. (1992) Dev. Brain Res., 67, 153-160). In the current study we extend our analysis of the changes in Purkinje cell numbers up to 24 months of age in +/sg and C57BL/6J wild-type mice. At 13 and 18 months, while wild-type Purkinje cell numbers remain unchanged, there is a 22-26% loss in the number of Purkinje cells in +/sg after which no further cell loss is observed. Between 18 and 24 months, however, a 22% loss of Purkinje cell occurs in +/+ animals, with the result that by 2 years of age, the size of the Purkinje cell population is again similar in both genotypes. Analysis of the cell loss in both the mediolateral and the anteroposterior dimensions, as well as the immunostaining of Purkinje cells in frontal sections, reveal no obvious regional variation in the Purkinje cell loss. These results suggest that in +/sg, a precocious process of aging affects the size of the Purkinje cell population.

mGluR1 mutant mice are viable but show characteristic cerebellar symptoms such as ataxic gait and intention tremor. The anatomy of the cerebellum is not overtly disturbed. Excitatory synaptic transmission from parallel fibers (PFs) to Purkinje cells and that from climbing fibers (CFs) to Purkinje cells appear to be functional, and voltage-gated Ca2+ channels of Purkinje cells are normal. Both PF and CF synapses display normal short-term synaptic plasticity to paired stimuli. By marked contrast, long-term depression (LTD) is clearly deficient and conditioned eyeblink response is impaired. We conclude that mGluR1 is required for the induction of LTD and that the ataxic behavior and impaired eyeblink conditioning of the mGluR1 mutant mice are primarily due to deficient LTD.

Cortical neurons are organized into highly patterned ensembles of cells, each with a distinct physiological function. The precursors of these cells arise in the ventricular zone, itself a highly patterned mosaic of cells. To determine whether the ventricular pattern presages the adult, investigators examine the process of cell migration. Recent evidence suggests that both radial (pattern-preserving) and tangential (pattern-blurring) cell movements occur during development.

The distribution of Purkinje cells, granule cells, and olivary neurons was quantitatively analyzed in a lurcher +/Lc in equilibrium C3H/HeJ chimera in which the surviving wild type Purkinje cells were unilaterally distributed in the left hemicerebella. The left hemisphere of this mouse contains 7600 Purkinje cells, approximately 10% of the number of Purkinje cells in inbred C3H/HeJ mice. The right hemisphere contains 300 Purkinje cells, all of which are found within 200 microns of the midline. As in other +/Lc in equilibrium wild type chimeras, the ratio of granule cells to Purkinje cells is increased in the left hemisphere, reflecting increased granule cell survival. In the right hemisphere, however, the number of granule cells is reduced to that found in +/Lc mutants. In the inferior olive, almost twice as many neurons are found in the right nucleus as opposed to the left nucleus. As the projections of olivary neurons are crossed, the number of olivary neurons is increased in the nuclei that project to the cerebellar hemisphere containing Purkinje cells compared to the olivary nuclei that project to the cerebellar hemisphere with almost no Purkinje cells. The preferential survival of granule cells and olivary neurons that either occupy or project to the hemicerebellum containing Purkinje cells suggests that the availability of trophic support from target Purkinje cell neurons is spatially restricted.

Introduction Massive neuronal loss is the prominent pathological hallmark feature of Alzheimer’s disease (AD). One of the ways post-mitotic neurons die is via their ectopic entry into cell-cycle, which often occurs at very early stages of the disease [1] . We have previously demonstrated occurrence of neuronal cell cycle events (CCEs) in a transgenic mouse model of AD (R1.40) similar to those occur in human AD [1] . Furthermore, these events were dependent upon amyloidogenic processing of amyloid-precursor protein [2] , precede deposition of senile plaques and coincide with elevated neuroinflammation. While we have also demonstrated that Tool-Like Receptor-4 (TLR4) mediated induction of neuroinflammation accelerated neuronal CCEs, and blocking inflammation via NSAIDs completely blocked neuronal CCEs and reduced microglial activation [3] , the source of neuroinflammation and molecular mechanisms for the induction of neuronal CCEs is unclear. Methods We utilized following methods: immunohistochemistry, purification and adoptive transfer of microglia, primary neurons and microglial cultures and generation and analysis of R1.40- Tnf α −/− mice. Results Here we demonstrate that neuroinflammation, cell autonomous to microglia, is capable of inducing neuronal CCEs. First, CD45+ microglia is present at six months of age in the affected region of R1.40 mouse brain and increases with age. Second, the F4/80+ microglia spatially co-exist with cyclin D1+ neurons in the temporal cortex of human AD brain. Notably, these cyclin D1+ neurons in the human brain are also TUNEL positive, suggesting that a portion of them is apoptotic. Third, purification and adoptive transfer of CD11b+ microglia derived form R1.40, but not from non-transgenic mice, was sufficient to induce cyclin D1 expression within the neurons of non-transgenic recipient mouse brain. Fourth, conditioned media (CM) derived from amyloid beta activated primary microglia significantly induced dose-dependent increase in DNA replication within primary cortical neurons and strongly correlated with increased levels of tumor-necrosis factor-α(TNFα) in the CM. Notably, blocking or activating TNFα pathway either via anti-TNFα antibody or recombinant human TNFα, significantly induced cyclin D1 expression and/or DNA replication in in vitro as well as in adoptive transfer studies. Generation of R1.40 mice deficient for TNFα (R1.40- Tnfa −/− ) displayed decreased microglial activation and showed significantly lower levels of cyclin D1+ neurons at six months of age. Finally, analysis of downstream molecules of TNFα receptor pathway suggested that neuronal CCEs is mediated via activation of JNK signaling pathway. Conclusion Together, our results provide first direct evidence that the neuroinflammation, cell-autonomous to microglia, induces AD-related neuronal CCEs and neurodegeneration via alteration of TNFR-MAPK signaling pathway.

Considerable evidence points to important roles for inflammation in Alzheimer's disease (AD) pathophysiology. Epidemiological studies have suggested that long-term nonsteroidal anti-inflammatory drug (NSAID) therapy reduces the risk for Alzheimer's disease; however, the mechanism remains unknown. We report that a 9-month treatment of aged R1.40 mice resulted in 90% decrease in plaque burden and a similar reduction in microglial activation. Ibuprofen treatment reduced levels of lipid peroxidation, tyrosine nitration, and protein oxidation, demonstrating a dramatic effect on oxidative damage in vivo. Fibrillar β-amyloid (Aβ) stimulation has previously been demonstrated to induce the assembly and activation of the microglial nicotinamide adenine dinucleotide phosphate (NADPH) oxidase leading to superoxide production through a tyrosine kinase-based signaling cascade. Ibuprofen treatment of microglia or monocytes with racemic or S-ibuprofen inhibited Aβ-stimulated Vav tyrosine phosphorylation, NADPH oxidase assembly, and superoxide production. Interestingly, Aβ-stimulated Vav phosphorylation was not inhibited by COX inhibitors. These findings suggest that ibuprofen acts independently of cyclooxygenase COX inhibition to disrupt signaling cascades leading to microglial NADPH oxidase (NOX2) activation, preventing oxidative damage and enhancing plaque clearance in the brain.

Contemplating Alzheimer's disease as cancer: a loss of cell-cycle controlThe nerve cells of the adult brain are born predominantly during embryogenesis, and as development proceeds, the addition of new nerve cells slows to a barely detectable trickle. By the time a human being nears the end of life most of their nerve cells have been in a post-mitotic state for seven to nine decades. The means by which cells accomplish this feat of suspended animation are unknown, but there are no apparent failures of the cell cycle arrest mechanism.During the past few years several laboratories have suggested that if a nerve cell violates the prohibition against cell division it will die rather than divide. The principle was first suggested when a powerful oncogene was expressed specifically in maturing Purkinje cells in transgenic mice. Unexpectedly, transgenic Purkinje cells began replicating their DNA but subsequently died 1xIn vivo viability of postmitotic Purkinje neurons requires pRb family member function. Feddersen, R.M. et al. Mol. Cell. Neurosci. 1995; 6: 153–167Crossref | PubMed | Scopus (58)See all References1. Next, researchers found that the activation of cell cycle processes is part of the mechanism by which developmental failure of trophic support leads to nerve cell death 2xThe induction of multiple cell cycle events precedes target-related neuronal death. Herrup, K. and Busser, J.C. Development. 1995; 121: 2385–2395PubMedSee all References, 3xAnalysis of cell cycle-related gene expression in postmitotic neurons: selective induction of Cyclin D1 during programmed cell death. Freeman, R. et al. Neuron. 1994; 12: 343–355Abstract | Full Text PDF | PubMed | Scopus (499)See all References.Now, several laboratories have explored the role of cell cycle processes in late-onset neurodegenerative disease. Our laboratory was one of several that provided immunocytochemical evidence for the presence of cell cycle activators (Fig. 1Fig. 1) as well as inhibitors in neurons of Alzheimer's disease brain. But the cell cycle is a highly regulated phenomenon.Fig. 1Immunocytochemical evidence for the re-expression of proliferating cell nuclear antigen (PCNA), a subunit of DNA polymerase. (a) Although normally found only in dividing cells, hippocampal neurons in Alzheimer's disease brain show occasional cells with significant expression of this protein. (b) Cells from a similar region of an age-matched, non-demented control show no evidence of expression.View Large Image | Download PowerPoint SlideOur laboratory adapted cytogenetics techniques such as fluorescent in situ hybridization (FISH) to analyze the DNA directly. We discovered that the chromosome complement in 4% of the hippocampal pyramidal cells in Alzheimer's disease has doubled 4xMice deficient for Rb are nonviable and show defects in neurogenesis and haematopoiesis. Yang, Y. et al. J. Neurosci. 2001; 21: 2661–2668PubMedSee all References4. Using either short centromeric FISH probes or long probes to unique sequences we were able to show that these neurons had up to four copies of each locus instead of the expected two – direct evidence that a coordinated DNA duplication had occurred (Fig. 2Fig. 2). No evidence of mitosis was found, suggesting the cells arrested at or near the G2 phase of the cell cycle.Fig. 2Fluorescence in situ hybridization for unique sites in the human genome. Neurons of the adult brain should all be diploid and display no more than two sites of hybridization. (a) In the Alzheimer's disease hippocampal neuron illustrated, four spots are found (white arrows) indicating that cells have arrested in G-phase, and that coordinated DNA duplication has occurred. (b) Cells from control subjects always have two spots or fewer.View Large Image | Download PowerPoint SlideThe implication of these findings is that the neurons in the Alzheimer's disease brain are subjected to a mitotic pressure that drives them to attempt a cell cycle they cannot complete. In most cancers, a gene mutation disrupts cell cycle suppression leading to the clonal expansion of a single dysregulated cell. If our model is correct, then the problem in Alzheimer's disease is nearly the exact opposite: although the nerve cells themselves remain genetically intact, extrinsic factors that promote cell division increase until, one by one, the most susceptible neurons divide and die.

Retrograde axonal transport of125I labeled nerve growth factor (NGF) occurs in the adrenergic neurons of the sympathetic nervous system of rats as well as mice. This transport is saturable, it can be blocked by an excess of unlabeled NGF and shows a specificity for the biologically active form of the protein. Other proteins such as serum albumin, chymotrypsin or insulin are transported in varying, but much smaller, amounts. Although the transport of albumin is blocked by an excess of NGF, the opposite is not true, again suggesting a specific mechanism. NGF antibody, injected subcutaneously, prevents retrograde transport in neonatal but not in more mature animals and the relative resistance of the latter to the action of NGF antiserum may be related to this phenomenon.

Genetically mosaic animals are valuable tools in the study of cell lineage relationships during development. The work described here is a quantitative analysis of large motor neurons of the facial nerve nucleus of the mouse. Aggregation chimeric mice were made from embryos which differed at the structural locus for the enzyme beta-glucuronidase. This difference is an intrinsic property of each cell and is easily detected by histochemical means. The numbers of cells in each of 10 facial nuclei from six glucuronidase chimeras were determined and each cell was scored for its embryo of origin using the glucuronidase marker. Unlike cerebellar Purkinje cells, this analysis revealed no evidence of numerical quanta of invariant size in the facial nucleus. Determination of the fraction of the total cells comprised by the descendants of each embryo, however, revealed evidence of quanta. These values suggest that the neurons of each facial nucleus descend from 12 progenitors. In agreement with similar calculations performed on Purkinje cells, the facial nucleus progenitors appear to be selected as the sole source of the facial nucleus neurons some time during the neural plate to neural groove stage of development. The absence of rigidly specified numerical quanta in the nucleus suggests that factors extrinsic to these cells contribute to the final adult number of neurons.

Leaner is an autosomal recessive mutation of the mouse which results in a severe ataxia accompanied by cellular losses in the cerebellar cortex. The purpose of this report is to construct a developmental profile of these losses. Of the three cerebellar cell types studied, the granule cells are the first to show obvious degenerative changes. Pycnotic cells are numerous in the internal granule cell layer at postnatal day 10, and, while they are found throughout the cortex, they are more concentrated in the anterior folia. Initially, there is a strong tendency for the pycnotic cells to be located in the deep half of the internal granule cell layer. By four postnatal months the rate of loss has slowed but the finding of occasional pycnotic cells in animals up to one year old suggests it continues for the life of the animal. Quantitative analysis of Purkinje and Golgi cells in leaner cerebella reveals a progressive loss of these cells as well. The number of Golgi cells falls uniformly to around half of the wild-type number. By contrast, the Purkinje cells show much more extensive degeneration. Further, the rate of cell death shows a regional variation; it is significantly more rapid in anterior folia. Overall, the number of Purkinje cells in leaner falls to about one-fifth of the wild-type number. The loss of both Purkinje and Golgi cells occurs late relative to the major events of cerebellar maturation. Significant cell loss is not observed until the end of the first postnatal month. For the next 4 to 6 weeks there is extensive cell death but, rather than abating, it appears to continue at a low rate for the life of the animal. It is hoped that this developmental sketch of the leaner defect will stimulate others to approach leaner and its alleles, tottering and rolling, as models for heterogeneity of disease expression.

The site of gene action of the Staggerer mutation of mice was investigated with Staggerer in equilibrium or formed from wild-type chimeras. Homozygous Staggerer mice show severe locomotor difficulties due to cerebellar abnormalities which include degeneration of virtually all granule cells and cytological defects in Purkinje cells. Although the locomotor deficits of the mutant were not present in the chimeras, the presence of Staggerer cells affected cerebellar structure. The size and the extent of foliation of the chimeric cerebella were intermediate between wile-type and homozygous Staggerer. A normally proportioned granule cell layer was present. Using beta-glucuronidase as an independent determinant of a cell's genotype, it was found that the genotypically Staggerer medium-to-large neurons expressed all of the light microscopic defects observable in these cells in the homozygous mutant. These defects include: (1) smaller size; (2) usually ectopic location; and (3) regional variation in the cytological appearance of the perikaryon. By contrast, all Purkinje cells which were genotypically wild-type appeared normal in size, in location and in their cytological appearance. Their density, however, was much reduced from wild-type. The effects of the Staggerer mutation on the granule, stellate and basket cells could not be directly assessed as the glucuronidase marker is not suitable for use with these cells. The Staggerer gene thus acts directly on Purkinje cells rather than via extracellular environmental changes. The findings are discussed in terms of their implications for normal cerebellar development.

The interaction of nerve growth factor (NGF) with its receptor on cells of the PC 12 cell line was studied. All experiments were done at 0.5 °C to minimize degradation and processes requiring membrane mobility. Under these conditions, a single class of high affinity binding sites with a dissociation constant of 2.9 × 10 −9 M was observed. The number of receptors per cell was 58000. The binding was linear with the number of cells in the assay and was not displaced by proteins other than native nerve growth factor. Trypsin treatment of the cells destroyed the specific binding. The removal of divalent cations had no effect on the binding. Culturing the cells for 2 weeks in NGF prior to assay did not change the receptor number or receptor affinity and there was a similar lack of effect of the density of the culture from which the cells were taken for assay. The present findings are compared with previous studies on the dorsal root ganglia and sympathetic ganglia neurons, and the implication for the use of PC 12 as a model for the study of NGF action are discussed.

This paper examines the structure and cytoarchitecture of the cerebellum of the weaver mutant mouse with particular emphasis on regional differences along the mediolateral and anterior-posterior axes. We have uncovered several, previously undescribed features of the weaver cerebellar phenotype. Perhaps the most dramatic example of our findings is the severe disruption of the folial structure of the hemispheres of the weaver cerebellum. A dorsal overgrowth of tissue occurs in the hemispheres that forms a finger-like projection superficial to an atrophie but structurally more normal cerebellar mass underneath. While this folial abnormality is most evident in the homozygote ( wv/wv ) the antecedents of its appearance are already apparent in the heterozygote ( +/wv ). At the level of the cytoarchitectonics of the mutant brain, we find substantial variation in the positioning, numbers and density of both Purkinje and granule cells. As a whole, Purkinje plus Golgi II cell numbers are down by over 40%, but this reduction occurs almost exclusively in the medial half of the cerebellum. The hemispheric region contains a nearly normal number of cells per sagittal section (although their positions are predominantly incorrect). The granule cells also show numerical variation; they are nearly absent at the midline, but a substantial number of them survive in the lateral cerebellar cortex. In the paraflocculus, for example, granule cells can be observed in a modest internal granule cell layer as late as 38 postnatal days. These results are discussed in terms of a model of wv gene action in which we propose that the effect of the mutation is a general disruption of cellular distribution in the cerebellar cortex, affecting both Purkinje and granule cells and beginning prenatally.

Staggerer (sg) is a mutation in mice which causes severe cerebellar atrophy. Virtually all of the granule cells degenerate by the end of the first postnatal month. The Purkinje cells are also affected. They are reduced in number, ectopic in location, reduced in size and show a regional variation in their cytological appearance. The appearance of the cerebellar cortex of Staggerer in equilibrium wild-type chimeric mice has demonstrated that the reduced size, ectopia, and regional variation are always traits of cells whose nucleus is of sg/sg genotype and thus are due to factors intrinsic to the affected cell. This report is a quantitative analysis of the cerebellar cortex of two Staggerer chimeras. Sections were chosen from serially sectioned half cerebella. Using beta-glucuronidase activity as an independent marker of a cell's genotype, cells were scored as belonging to 1 of 3 cell classes: wild-type Purkinje cell, wild-type Golgi cell, and Staggerer medium-to-large neuron. The results confirm that the total number of cells in the chimera (all 3 classes) is greater than that found in a homozygous Staggerer but less than that found in a wild-type. Further, the distribution of the two genotypes strongly suggests that the decreased number of cells in Staggerer and in the chimera is due to an intrinsic defect in the Staggerer cells and not to a general (extrinsic) compromising of cerebellar development. Possible implications for normal cerebellar development are discussed.

Recent quantitative studies of lurcher chimeric mice have shown that the adult population of cerebellar Purkinje cells can properly be described as a small number of developmental clones of cells. The clones are not seen as patches of contiguous neurons; rather, the cells of any one clone distribute throughout the half-cerebellum that contains them, intermingling extensively with the Purkinje cells of other linkages. Lurcher ⇌ wild-type chimeras were analyzed using the cell autonomous Purkinje-cell-lethal mutant, lurcher ( + L ), as a cell marker. Cell counts from these chimeras revealed that the number of surviving Purkinje cells was always an integral multiple of a unit clone size. These numerical quanta are the evidence for the existence of Purkinje cell developmental clones. When two different inbred strains of mouse were compared (C3H/HeJ and C57BL/6), the resulting clonal analysis showed that the unit clone size (i.e., the number of Purkinje cells in one quantum) is an autonomous property of the lineage and hence, presumably, intrinsic to the progenitor cell that founded it. The current study uses the lurcher chimeric mouse system to examine the cell lineage relationships among the Purkinje cells of a third inbred strain of mouse, AKR/J. The data both support and extend our previous studies. Quantitative analysis reveals that the Purkinje cells of this strain also exist in clones, and the size of these clones is also strain-specific. The number of cells in a single clone (7850), however, is different from either C3H/HeJ (10,200) or C57BL/6 (9200). The fact that this value is so highly polymorphic among the inbred strains of mouse makes it likely that, rather than being a function of different alleles at a single genetic locus, clone size may well represent a multifactorial (but still cell-autonomous) property of developing Purkinje cells. Additional results from a single chimeric animal suggest strongly that clone number (i.e., the number of progenitors selected to found the population) is not strain-specific but results instead from cell:cell interactions during early central nervous system formation.

Quantitative morphological techniques have revealed several new aspects of the action of the Staggerer mutant gene. Staggerer is an autosomal recessive gene which causes ataxia and severe malformation of the cerebellar cortex in mice. The Purkinje cells of the cerebellar cortex are small, abnormal in morphology and reduced in numbers. The close synaptic and developmental relationship of Purkinje cells with the cells of the deep cerebellar nuclei (dcn) lead us to look for effects of the Staggerer mutation on the dcn neurons. The volume of the deep nuclear region is shrunken in Staggerer and there is a reduction in the volume of the white matter. These findings account for the reduced wet weights and protein concentration found by Roffler-Tarlov and Sidman. In contrast to the cells of the cortex, where 75% of the medium-to-large neurons are missing, the number of cells present in Staggerer dcn is identical to wild-type. The dcn neurons are not completely spared, however. Measurements of cross-sectional cell area revealed a 30% shrinkage of neurons in Staggerer dcn. The most likely interpretation of previous work and the current findings is that the Staggerer gene acts early in development but exerts its effects directly only on those derivatives of the ventricular zone in the roof of the fourth ventricle which are destined to become Purkinje and Golgi cells.

In the January 1984 issue of the Journal of Neuroscience 1 Angel de Blas presents a compelling analysis of the cytoarchitecture of the mammalian cerebellar cortex. While the major workers in this field have contributed the substantial groundwork on which this paper rests, de Blas brings to a well-studied brain region a fresh technical approach that emphasizes some potentially important relationships between neuron and glia. Two of the more fascinating questions addressed by this work are: (1) are the Bergmann glia of the cerebellar cortex involved in helping to establish the well known espalier shape of the Purkinje cell dendrite (flattened in the plane transverse to the long axis of the folium) and (2) do the Bergmann glia in any way serve to orient or guide the granule cell axons (i.e. the parallel fibers of the molecular layer) in a direction perpendicular to the plane of the Purkinje cell dendrite.

Retinoblastoma tumors appear to result from changes in a specific DNA sequencethat happen during a critical period of retinal development. These intraocular tumors are diagnosed in early childhood, usually from birth to four years of age. They are the apparent manifestation of a block in the differentiation of retinal cells, such that the cells keep dividing and may become malignant. Recent genetic studies have implicated a specific region of the human genome in the formation of retinoblastoma, and have elucidated some of the mechanisms by which a retinal cell can lose normal function in this region.

In a recent issue of Cell , Arthur Roach, Kevin Boylan, Suzanna Horvath, Stanley Prusiner, and Leroy Hood 1 reported their study of a cloned cDNA sequence representing the myelin basic protein of the rat. While reports of the isolation of cDNA clones are increasingly frequent, this paper represents a milestone in the emerging field of molecular neurogenetics. It is in some respects an answer to the lament of Barnstable et al. 2 that neurobiologists are ‘raising important questions that have no molecular answers' while molecular biologists have ‘arrived at answers for which they now have to discover the appropriate questions'. Roach et al. have succeeded in bridging this gap.