Alzheimer’s disease (AD) is the most common cause of dementia of the elderly. Although rare, familial AD is most often caused by one of only three known disease genes one of which is APP(amyloid precursor protein). APP is the source of the Aβ peptide fr...

In the mammalian visual system, three functionally distinct parallel processing streams extend from the retina to the visual thalamus and then to the visual cortex: magnocellular (M), parvocellular (P), and koniocellular (K). Tree shrews ( Tupaia belangeri ), a preprimate species, provide an advantageous model to study the K pathway in isolation because, while M and P pathways remain mixed in Lamina 1 (L1), L2, L4, and L5 of the lateral geniculate nucleus (LGN), L3 and L6 receive strictly K-input from the contralateral eye. Additionally, K-input laminae selectively receive glutamatergic axons from the superior colliculus. To reveal how cellular and synaptic properties of K geniculate laminae may differ from M/P laminae and how tectal input may shape the K relay to the cortex, we studied the morphology and connectivity of retinal and tectal terminals in pathway-specific laminae. While confirming that K laminae relay cells contain calbindin, we also found its expression in GABAergic cells across all laminae....

Coordinated activation of integrin and growth factor signaling is necessary for efficient axon elongation induced by NGF. However, exactly where neurotrophin and integrin signaling pathways intersect in the mediation of axon growth is not known. Our work and the work of others have shown that inhibition of integrin-linked kinase (ILK), a β1 and β3 integrin subunit bound serine/threonine kinase, blocks NGF-induced axon growth on laminin by reducing GSK-3β phosphorylation (Mills et al., J. Neurosci. 23(5): 1638-48, 2003; Zhou et al. Neuron, in press). We show here that embryonic dorsal root ganglion (DRG) neurons plated on polylysine in the absence of integrin activation also show reduced GSK-3β phosphorylation and decreased axon growth in response to NGF compared to embryonic DRG neurons plated on laminin. Together these data suggest that integrin and NGF signaling may converge through ILK to regulate GSK-3β activity and axon growth. Furthermore, by using a variety of kinase inhibitors, we have identified a...

Electrophysiological recording followed by BDA microiontophoretic injections in neurons of the primary motor cortex (M1) of cynomolgus monkeys (Macaca fascicularis) and in the centre median/parafascicular thalamic complex (CM/Pf) of squirrel monkeys (Saimiri sciureus) have revealed the following facts. (1) Thalamostriatal projections. The majority of individually labeled axons that emerged from CM/Pf complex in squirrel monkeys innervated both motor cortex and striatum. These axons emitted, in some cases, a few collaterals in the reticular thalamic nucleus and bifurcated within the internal capsule. One branch ascended via the corona radiata to the cerebral cortex, where it arborized moderately in layers V and VI, and much less profusely in layer I. The other branch entered the striatum, where it broke out into 3-5 smaller collaterals. These thin branches divided into numerous shorter collaterals that displayed pedunculated varicosities organized as dense, cluster-like terminal fields. Axons from the CM an...

One strategy used by the developing nervous system to ensure the establishment of appropriate connectivity is to overproduce axon collaterals and then to select only those that make correct connections. This is thought to be largely driven by activity and trophic factor-dependent axon competition. Here, we have asked how activity and one class of trophic factors, the neurotrophins, regulate this process of axon collateral selection using cultured sympathetic neurons in compartmented cultures as a model system. We show that in the presence of NGF, local depolarization confers a growth advantage on depolarized axon collaterals, acting both to enhance the rate of growth and to increase axonal density. This growth advantage occurs at the expense of unstimulated collaterals deriving from the same neurons which exhibit reduced axonal growth even when compared to axons from control neurons growing into the same amount of NGF. Depolarization mediates the enhanced growth of the stimulated axons by locally activatin...

As part of a series of feasibility studies on culture of cryopreserved dissociated neurons from varied regions of the nervous system we report on cryopreservation and subsequent functional studies on dorsal root ganglion cells and their axonal outgrowth. Rat embryonic dorsal root ganglion cells were isolated and dissociated using standard procedures. A method was developed to cryopreserve aliquots of 200 K DRG neurons which could be subsequently thawed and plated at 5 K per well (PDL coated 96 well plates) and grown in Neurobasal medium with B27. No further supplementation was required for survival to 19+ days in vitro. In culture, cryopreserved DRG neurons showed the same size and neurochemical type distribution as freshly dissociated dorsal root ganglion neurons. Over the first few days neuronal cell bodies formed ganglionated clusters and fasiculated axon bundles, but with the addition of mitotic inhibitors DRG neurons and their axons remain dispersed. Cells were transfected with eGFP using Transmesseng...

The myelin sheath enables saltatory conduction by demarcating the axon into a small nodal region for excitation and an extended, insulated internodal region for efficient spread of passive current. This anatomical demarcation produces a dramatic heterogeneity in ionic fluxes during excitation, a classical example being the restriction of Na influx at the node. However, ionic fluxes are not restricted only to Na during excitation, as recent studies have revealed that action potentials also induce calcium influx into axons of mammalian optic nerves. Does calcium influx along a myelinated axon also show spatial heterogeneity during excitation, as in the case for sodium? To address this, we analyzed spatial profiles of axonal calcium transients during excitation by selectively staining axons with calcium indicators, and analyze the data by two models, one assuming uniform axonal influx and the other assuming discrete nodal influx, with a coefficient of axial calcium diffusion empirically determined in situ usi...

To establish the intricate neuronal network with necessary precision and reliability, one essential process is that neurons must correctly project their axons to their synaptic targets during embryonic development. In vivo, axonal tips (“growth cones”) navigate through a three-dimensional (3-D) environment, and the factors that can steer growth cones include ECM adhesion molecules, topographic features of the environment, and soluble chemotropic molecules. However, in most in vitro studies, neurons were confined on a uniform two-dimensional substrate, hence it was difficult to investigate the integration process in the presence of multiple contact cues. We have studied the growth cone 3-D navigation and axon decision-making using structured polydimethylsiloxane (PDMS) substrates combined with Matrigel. PDMS substrates with grooves/wells of various width and depth were fabricated by standard photolithography and soft lithography procedures, then coated with poly-D-lysine or laminin. Murine embryonic cortica...

Subjects with schizophrenia exhibit both impairments in working memory and deficits in markers of GABA neurotransmission in the prefrontal cortex (PFC). The coordinated activity of specific populations of pyramidal neurons and GABA-containing cells in the primate PFC appears to be critical for working memory. Different subclasses of GABA-containing cells can be distinguished by their content of the calcium-binding proteins parvalbumin (PV) or calretinin (CR). Knowledge of the synaptic targets of the CR- and PV-containing cells is important to understanding their role in the PFC circuitry involved in working memory. The postsynaptic targets of PV-containing cells have been well-characterized in many cortical areas, including the PFC. However, the targets of CR-containing cells have only been examined in visual cortex, where their synaptic targets differ by layer. Thus, in the present study, we used immuno-electron microscopy to examine the synaptic targets of CR-labeled axon terminals in the superficial and...

The cellular prion protein (PrPC) plays a crucial role in transmissible spongiform encephalopathies (TSEs). PrPC and is predominanatly expressed in brain, at neuronal synapses, axons and cell bodies. It is attached by a glycosyl-phosphatidyl inositol (GPI) anchor to the outer cell surface and its physiological function is still elusive. PrP knockout mice (prnp-/-), in which only the PrP coding sequence was disrupted, develop normally and remain healthy after birth. Several lines of evidence suggest a role of PrPC in neurite outgrowth. We have investigated the effect of PrPC expression on survival and axon growth using motoneurons from prnp-/-, prnp+/- and wild-type mice. These motoneurons were cultured with BDNF, CNTF or GDNF as a neurotrophic factor. No reduction in cell survival was observed but axon length was significantly shorter in comparison to heterozygote or wild-type controls. The reduction of axon length occured independently of the neurotrophic factor applied, and with both laminin or laminin-2...