Makoto Nishiyama, M.D., Ph.D.

Theoretical modeling for axon guidance and polarization
Honda N.; Nishiyama M.; Hong K.; Ishii S.
2011 ;71:e11-e11, Neuroscience research
The unique capability of the growth cone is that it can show bi-directional response (attraction and repulsion) to the same cue molecules depending on the biological situations such as expressed receptors and adhesion molecules, although most of other chemotactic cells only show uni-directional response (attraction or repulsion) to their specific cue molecules.Wehave experimentally clarified that the bi-directional response of the growth cone against netrin-1 is regulated by the intracellular level of cAMP and cGMP so that the direction is determined by the ratio of these signals; as the ratio increased, turning response was continuously changed from repulsion to attraction. To elucidate the mechanism underlying the bi-directional gradient sensing, we developed a theoretical model based on reaction-diffusion system consisting of activator and inhibitor molecules. Through mathematical analyses, we found that bi-directional responses can be produced, controlled by the ratio of activator to inhibitor signals, which is consistent with the experimental observation. Also, our analysis characterized both uni- and bi-directional responses from the point of view of adaptation. In polarization, a neuron initially has several neurites, but only one elongates and develops into an axon. This process is robustly controlled; when multiple neurites are selected, the selection is eventually reduced to yield a single axon.Whatis the system by which morphological information is decoded differently based on the presence of a single or multiple axons? To answer this question, we constructed a biophysical model with the active transport. Our mathematical analysis revealed that, as neurites elongate, transported factors accumulate in the growth cone but are degraded during diffusion to the soma. Such a system effectively works as local activation-global inhibition mechanism, resulting in robust polarization
— id: 137092, year: 2011, vol: 71, page: e11, stat: Journal Article,

Semaphorin 3A induces Ca(V)2.3 channel-dependent conversion of axons to dendrites
Nishiyama, Makoto; Togashi, Kazunobu; von Schimmelmann, Melanie J; Lim, Chae-Seok; Maeda, Shin-Ichi; Yamashita, Naoya; Goshima, Yoshio; Ishii, Shin; Hong, Kyonsoo
2011 Jun;13(6):677-686, Nature cell biology
Polarized neurites (axons and dendrites) form the functional circuitry of the nervous system. Secreted guidance cues often control the polarity of neuron migration and neurite outgrowth by regulating ion channels. Here, we show that secreted semaphorin 3A (Sema3A) induces the neurite identity of Xenopus spinal commissural interneurons (xSCINs) by activating Ca(V)2.3 channels (Ca(V)2.3). Sema3A treatment converted the identity of axons of cultured xSCINs to that of dendrites by recruiting functional Ca(V)2.3. Inhibition of Sema3A signalling prevented both the expression of Ca(V)2.3 and acquisition of the dendrite identity, and inhibition of Ca(V)2.3 function resulted in multiple axon-like neurites of xSCINs in the spinal cord. Furthermore, Sema3A-triggered cGMP production and PKG activity induced, respectively, the expression of functional Ca(V)2.3 and the dendrite identity. These results reveal a mechanism by which a guidance cue controls the identity of neurites during nervous system development
— id: 133465, year: 2011, vol: 13, page: 677, stat: Journal Article,

Identification of a molecular system that regulates growth cone membrane potential during growth cone guidance
Yamada T.; Jimbo H.C.; Ishii S.; Nishiyama M.; Hong K.; Sakumura Y.
2011 ;12:?-?, BMC Neuroscience
Growth cones, at the tips of growing neurites (axons and dendrites) of developing neurons are guided to their synaptic targets by various external guidance molecules to make functional neuronal connections. The mechanisms by which growth cones respond to these external (Figure presented) guidance molecules are yet to be elucidated. In cultured Xenopus spinal commissural interneurons (CINs) derived from stage 26-28 embryos, the magnitude of cyclic-GMP (cGMP) signaling determines the direction of growth cone turning in response to a secreted guidance molecule, semaphorin 3A (Sema3A) by controlling growth cone membrane potential shifts [1]. The molecular mechanism of intracellular signaling that controls the growth cone membrane potential is largely unknown. In this study, we aim to identify the molecular interaction between the signaling cascade(s) that regulate(s) the activities of Na⁺ channels (NaC) and Cl- channels (ClC) by employing Bayesian statistics with parametric models for the computational analysis of experimentally derived data. Measurements of membrane potential in cultured Xenopus spinal CIN growth cones were performed in the absence (control) and presence of the following pharmacological drugs: DNDS (ClC blocker), STX (NaC blocker) and KT5823 (PKG inhibitor) (Fig. 1). The following conjectures were applied in the model: 1) Both NaC and ClC are regulators of growth cone membrane potential shifts in the control. 2) In the presence of either DNDS or STX, one of the ionic channels does not function without changing the intracellular molecular system. 3) In the presence of KT5823, the molecular system is modified by inhibition of the PKG-mediated signaling cascade, which is required for activation of the NaC. Under these assumptions, if an interaction occurs between the cascades that regulate NaC and ClC, the time courses of membrane potential shifts in the presence of STX should be different from those in the presence of KT5823. However, a problem exists in modeling because of the variability in the cellular response to a given cGMP stimulus (Fig. 1) that maybe due to various cellular characteristics, e.g., growth cone shape, channel distribution, and developmental stage of the animal. Thus, using a statistical characterization of the molecular system, we examined the existence of such an interaction based on the ability to reconstruct the time courses in the control condition. We reconstructed time courses of membrane potential shifts from the data set of DNDS/STX or DNDS/KT5823. We introduced the Hill-type formula as a parametric expression of the time course since most feedforward biochemical signals show a monotonic increase. The cellular variability in this framework can be modeled as the probability distributions of the parameters, which is approximated with the Gamma distributions. We estimated the optimized hyper-parameters in the Gamma distributions for all pharmacological conditions in which the posterior distributions are maximized. The difference in the hyperparameters between the STX condition and the KT5823 condition suggests that there exists an interaction between the cascades that regulate the NaC and ClC
— id: 146281, year: 2011, vol: 12, page: ?, stat: Journal Article,

From guidance signals to movement: signaling molecules governing growth cone turning
Hong, Kyonsoo; Nishiyama, Makoto
2010 Feb;16(1):65-78, Neuroscientist
Directed growth cone movements in response to external guidance signals are required for the establishment of functional neuronal connections during development, adult nerve regeneration, and adult neurogenesis. Growth cone intrinsic properties permit different growth cone responses (e.g., attraction or repulsion) to a guidance signal, and alterations to these intrinsic properties often result in opposite growth cone responses. This article reviews the current knowledge of growth cone signaling, emphasizing the dependency of Ca(2+) signaling on membrane potential shifts, and cyclic nucleotide and phosphoinositide signaling pathways during growth cone turning in response to guidance signals. We also discuss how asymmetrical growth cone signaling is achieved for the fine-tuned growth cone movement
— id: 111638, year: 2010, vol: 16, page: 65, stat: Journal Article,

Semaphorin 3A specifies dendrites from axons through the activation of CaV2.3 channels
Nishiyama M.; Togashi K.; Von Schimmelmann M.J.; Goshima Y.; Hong K.
2010 ;68:e62-e62, Neuroscience research
Ion channels present in membranes of all cells, mediate cell communication, cellular homeostasis and polarized cellular responses. Here we report that a secreted semaphorin 3A, Sema3A, controls neurite polarization - axon/dendrite specification - of Xenopus spinal neurons via activation of R-type Ca²⁺ (CaV2.3) channels. In cultured Xenopus spinal neuron growth cones, Sema3A induces R-type Ca²⁺ currents, but suppresses L- and N-type Ca²⁺ currents; Sema3A also suppresses axon specification, which is reversed by CaV2.3 channel blocking. In tadpole spinal cords, down-regulation of Sema3A or its receptor neuropilin-1 (Npn-1) expression inhibits CaV2.3 alpha1E expression and dendrite formation. Elimination of functional CaV2.3 channels prevents dendrite formation while increasing axon specification. This failure of dendrite formation is reversed by ectopic expression of the mouse CaV2.3 alpha1E subunit, but only when functional Sema3A is present. Our studies therefore demonstrate that Sema3A not only signals to guide growing axons but also signals to control axon/dendrite fates by concomitantly suppressing axon specification and promoting dendrite formation through the functional CaV2.3 channel expression
— id: 113672, year: 2010, vol: 68, page: e62, stat: Journal Article,

GABAergic activities control spike timing- and frequency-dependent long-term depression at hippocampal excitatory synapses
Nishiyama, Makoto; Togashi, Kazunobu; Aihara, Takeshi; Hong, Kyonsoo
2010 ;2:22-22, Frontiers in synaptic neuroscience
GABAergic interneuronal network activities in the hippocampus control a variety of neural functions, including learning and memory, by regulating theta and gamma oscillations. How these GABAergic activities at pre- and postsynaptic sites of hippocampal CA1 pyramidal cells differentially contribute to synaptic function and plasticity during their repetitive pre- and postsynaptic spiking at theta and gamma oscillations is largely unknown. We show here that activities mediated by postsynaptic GABA(A)Rs and presynaptic GABA(B)Rs determine, respectively, the spike timing- and frequency-dependence of activity-induced synaptic modifications at Schaffer collateral-CA1 excitatory synapses. We demonstrate that both feedforward and feedback GABA(A)R-mediated inhibition in the postsynaptic cell controls the spike timing-dependent long-term depression of excitatory inputs ('e-LTD') at the theta frequency. We also show that feedback postsynaptic inhibition specifically causes e-LTD of inputs that induce small postsynaptic currents (<70 pA) with LTP-timing, thus enforcing the requirement of cooperativity for induction of long-term potentiation at excitatory inputs ('e-LTP'). Furthermore, under spike-timing protocols that induce e-LTP and e-LTD at excitatory synapses, we observed parallel induction of LTP and LTD at inhibitory inputs ('i-LTP' and 'i-LTD') to the same postsynaptic cells. Finally, we show that presynaptic GABA(B)R-mediated inhibition plays a major role in the induction of frequency-dependent e-LTD at alpha and beta frequencies. These observations demonstrate the critical influence of GABAergic interneuronal network activities in regulating the spike timing- and frequency-dependences of long-term synaptic modifications in the hippocampus
— id: 128805, year: 2010, vol: 2, page: 22, stat: Journal Article,

Membrane potential shifts caused by diffusible guidance signals direct growth-cone turning
Nishiyama, Makoto; von Schimmelmann, Melanie J; Togashi, Kazunobu; Findley, William M; Hong, Kyonsoo
2008 Jul;11(7):762-771, Nature neuroscience
Plasma membrane potentials gate the ion channel conductance that controls external signal-induced neuronal functions. We found that diffusible guidance molecules caused membrane potential shifts that resulted in repulsion or attraction of Xenopus laevis spinal neuron growth cones. The repellents Sema3A and Slit2 caused hyperpolarization, and the attractants netrin-1 and BDNF caused depolarization. Clamping the growth-cone potential at the resting state prevented Sema3A-induced repulsion; depolarizing potentials converted the repulsion to attraction, whereas hyperpolarizing potentials had no effect. Sema3A increased the intracellular concentration of guanosine 3',5'-cyclic monophosphate ([cGMP]i) by soluble guanylyl cyclase, resulting in fast onset and long-lasting hyperpolarization. Pharmacological increase of [cGMP](i) caused protein kinase G (PKG)-mediated depolarization, switching Sema3A-induced repulsion to attraction. This bimodal switch required activation of either Cl(-) or Na+ channels, which, in turn, regulated the differential intracellular Ca2+ concentration increase across the growth cone. Thus, the polarity of growth-cone potential shifts imposes either attraction or repulsion, and Sema3A achieves this through cGMP signaling
— id: 86547, year: 2008, vol: 11, page: 762, stat: Journal Article,

Cyclic GMP-gated CNG channels function in Sema3A-induced growth cone repulsion
Togashi, Kazunobu; von Schimmelmann, Melanie J; Nishiyama, Makoto; Lim, Chae-Seok; Yoshida, Norihiro; Yun, Bokyoung; Molday, Robert S; Goshima, Yoshio; Hong, Kyonsoo
2008 Jun 12;58(5):694-707, Neuron
Cyclic nucleotide-gated channels (CNGCs) transduce external signals required for sensory processes, e.g., photoreception, olfaction, and taste. Nerve growth cone guidance by diffusible attractive and repulsive molecules is regulated by differential growth cone Ca2+ signaling. However, the Ca2+-conducting ion channels that transduce guidance molecule signals are largely unknown. We show that rod-type CNGC-like channels function in the repulsion of cultured Xenopus spinal neuron growth cones by Sema3A, which triggers the production of the cGMP that activates the Xenopus CNGA1 (xCNGA1) subunit-containing channels in interneurons. Downregulation of xCNGA1 or overexpression of a mutant xCNGA1 incapable of binding cGMP abolished CNG currents and converted growth cone repulsion to attraction in response to Sema3A. We also show that Ca2+ entry through xCNGCs is required to mediate the repulsive Sema3A signal. These studies extend our knowledge of the function of CNGCs by demonstrating their requirement for signal transduction in growth cone guidance
— id: 79567, year: 2008, vol: 58, page: 694, stat: Journal Article,

Cyclic AMP/GMP-dependent modulation of Ca2+ channels sets the polarity of nerve growth-cone turning
Nishiyama, Makoto; Hoshino, Akemi; Tsai, Lily; Henley, John R; Goshima, Yoshio; Tessier-Lavigne, Marc; Poo, Mu-Ming; Hong, Kyonsoo
2003 Jun 26;424(6943):990-995, Nature
Signalling by intracellular second messengers such as cyclic nucleotides and Ca2+ is known to regulate attractive and repulsive guidance of axons by extracellular factors. However, the mechanism of interaction among these second messengers in determining the polarity of the guidance response is largely unknown. Here, we report that the ratio of cyclic AMP to cyclic GMP activities sets the polarity of netrin-1-induced axon guidance: high ratios favour attraction, whereas low ratios favour repulsion. Whole-cell recordings of Ca2+ currents at Xenopus spinal neuron growth cones indicate that cyclic nucleotide signalling directly modulates the activity of L-type Ca2+ channels (LCCs) in axonal growth cones. Furthermore, cGMP signalling activated by an arachidonate 12-lipoxygenase metabolite suppresses LCC activity triggered by netrin-1, and is required for growth-cone repulsion mediated by the DCC-UNC5 receptor complex. By linking cAMP and cGMP signalling and modulation of Ca2+ channel activity in growth cones, these findings delineate an early membrane-associated event responsible for signal transduction during bi-directional axon guidance
— id: 39176, year: 2003, vol: 424, page: 990, stat: Journal Article,

Calcium signalling in the guidance of nerve growth by netrin-1
Hong K; Nishiyama M; Henley J; Tessier-Lavigne M; Poo M
2000 Jan 6;403(6765):93-98, Nature
Pathfinding by growing axons in the developing nervous system is guided by diffusible or bound factors that attract or repel the axonal growth cone. The cytoplasmic signalling mechanisms that trigger the responses of the growth cone to guidance factors are mostly unknown. Previous studies have shown that the level and temporal patterns of cytoplasmic Ca2+ can regulate the rate of growth-cone extension in vitro and in vivo. Here we report that Ca2+ also mediates the turning behaviour of the growth cones of cultured Xenopus neurons that are induced by an extracellular gradient of netrin-1, an established diffusible guidance factor in vivo. The netrin-1-induced turning response depends on Ca2+ influx through plasma membrane Ca2+ channels, as well as Ca2+-induced Ca2+ release from cytoplasmic stores. Reduction of Ca2+ signals by blocking either of these two Ca2+ sources converted the netrin-1-induced response from attraction to repulsion. Activation of Ca2+-induced Ca2+ release from internal stores with a gradient of ryanodine in the absence of netrin-1 was sufficient to trigger either attractive or repulsive responses, depending on the ryanodine concentration used. These results support the model that cytoplasmic Ca2+ signals mediate growth-cone guidance by netrin-1, and different patterns of Ca2+ elevation trigger attractive and repulsive turning responses
— id: 21642, year: 2000, vol: 403, page: 93, stat: Journal Article,

Calcium stores regulate the polarity and input specificity of synaptic modification
Nishiyama M; Hong K; Mikoshiba K; Poo MM; Kato K
2000 Nov 30;408(6812):584-588, Nature
Activity-induced synaptic modification is essential for the development and plasticity of the nervous system. Repetitive correlated activation of pre- and postsynaptic neurons can induce persistent enhancement or decrement of synaptic efficacy, commonly referred to as long-term potentiation or depression (LTP or LTD). An important unresolved issue is whether and to what extent LTP and LTD are restricted to the activated synapses. Here we show that, in the CA1 region of the hippocampus, reduction of postsynaptic calcium influx by partial blockade of NMDA (N-methyl-D-aspartate) receptors results in a conversion of LTP to LTD and a loss of input specificity normally associated with LTP, with LTD appearing at heterosynaptic inputs. The induction of LTD at homo- and heterosynaptic sites requires functional ryanodine receptors and inositol triphosphate (InsP3) receptors, respectively. Functional blockade or genetic deletion of type 1 InsP3 receptors led to a conversion of LTD to LTP and elimination of heterosynaptic LTD, whereas blocking ryanodine receptors eliminated only homosynaptic LTD. Thus, postsynaptic Ca2+, deriving from Ca2+ influx and differential release of Ca2+ from internal stores through ryanodine and InsP3 receptors, regulates both the polarity and input specificity of activity-induced synaptic modification
— id: 21643, year: 2000, vol: 408, page: 584, stat: Journal Article,

A ligand-gated association between cytoplasmic domains of UNC5 and DCC family receptors converts netrin-induced growth cone attraction to repulsion
Hong K; Hinck L; Nishiyama M; Poo MM; Tessier-Lavigne M; Stein E
1999 Jun 25;97(7):927-941, Cell
Netrins are bifunctional: they attract some axons and repel others. Netrin receptors of the Deleted in Colorectal Cancer (DCC) family are implicated in attraction and those of the UNC5 family in repulsion, but genetic evidence also suggests involvement of the DCC protein UNC-40 in some cases of repulsion. To test whether these proteins form a receptor complex for repulsion, we studied the attractive responses of Xenopus spinal axons to netrin-1, which are mediated by DCC. We show that attraction is converted to repulsion by expression of UNC5 proteins in these cells, that this repulsion requires DCC function, that the UNC5 cytoplasmic domain is sufficient to effect the conversion, and that repulsion can be initiated by netrin-1 binding to either UNC5 or DCC. The isolated cytoplasmic domains of DCC and UNC5 proteins interact directly, but this interaction is repressed in the context of the full-length proteins. We provide evidence that netrin-1 triggers the formation of a receptor complex of DCC and UNC5 proteins and simultaneously derepresses the interaction between their cytoplasmic domains, thereby converting DCC-mediated attraction to UNC5/DCC-mediated repulsion
— id: 21646, year: 1999, vol: 97, page: 927, stat: Journal Article,