To investigate whether also maintenance of the gradi- ent is activity-dependent, we analyzed mice with cochlear ablation after hearing-onset. To this end, NMRI mice were subjected to bilateral cochlea removal at P27, i.e. roughly 2 weeks after hearing-onset. Successfully operated animals were killed 3 weeks after deafening at P48. Subsequent anti- VIAAT immunohistochemistry revealed the persistence of the gradient in these mice (Fig. 5 A b, b′). There was a significant 18% upregulation of the immunosignal in the lateral limb compared to the medial one (Fig. 5 B; medial: 1.176 ± 0.115, lateral: 1.379 ± 0.122. p = 0.000242). This result indicates that activity deprivation after hearing- onset does not influence the VIAAT gradient, at least dur- ing a 3-week period. Taken together, these results reveal that neuronal activity is required for the formation of the VIAAT-ir gradient in the LSO, but not for its maintenance.
In particular, the members of the neurotrophin family are potential candidates for the above mentioned survival factor and have been implicated in mediating activity- dependent morphological plasticity. Especia lly brain-derived neurotrophic factor (BDNF) is known to be involved in LTP and LTD. The lack of BDNF, either as a result of gene or protein inactivation, leads to a profound inhibition of NMDAR-LTP (Korte et al., 1995; Patterson et al., 1996). In line with this, the addition of BDNF to hippocampal slices isolated from wildtype animals leads to a long- lasting enhancement of synaptic transmission (Kang and Schuman, 1995). Conversely, application of exogenous BDNF prevents LTD in the visual cortex (Kinoshita et al., 1999). Culturing dissociated hippocampal neurons in BDNF induces formation of synapses on these neurons (Vicario- Abejon et al., 1998). Overexpressing BDNF in slices from the visual cortex was found to lead to destabilization of spines resulting in increased growth and retraction rates (Horch et al., 1999) also indicating that BDNF is important for synapse (and spine) formation and maintenance.
Synaptic plasticity refers to the ability of the brain to modify its synaptic connectivity in an activity-dependent way. Its most prominent form was discovered in 1973 by Bliss and Lomo and is called long-term potentiation (LTP). Many of its properties make it ideally suited to serve as a cellular mechanism of learning and memory. In fact, a large body of data indicates that synaptic plasticity plays an important role in learning and memory formation. While much is known about the cell biological mechanisms of how changes in the strength of synaptic transmission are initiated, it remains unclear how these functional changes are laid down more permanently. A long-standing hypothesis in neuroscience posits that changes in neuronal morphology are a key mechanism for how information can be maintained in a synaptic network in an enduring way. In recent years, novel labeling and microscopic techniques have made it possible to image neuronal structure at the level of single synapses in time-lapse experiments. This approach has uncovered a wealth of information on how neuronal structure can change in response to synaptic activation. While many studies have focused on dendritic spines, which represent the postsynaptic structural part of most excitatory synapses in the brain, surprisingly little effort has gone into examining the structural plasticity of the presynaptic axon terminals that provide the synaptic input onto the dendritic spines. It is important not to leave out the presynaptic side, because presynaptic structural changes would be expected to have qualitatively very different consequences for the functional connectivity of the synaptic network. Moreover, examining both sides of the synapse at the same time is expected to yield important insights into how these critical compartments interact during plasticity.
Whilst plasticity in some areas of the CNS, like those associated with memory formation, reaches far into adulthood, the majority of areas lose their capacity to adapt after they have been fully established. In the cat visual system, for example, Hubel and Wiesel were the first to show that closing one eye and preventing it from exciting neurons during the first few months of life creates a visual cortex mainly representing the nondeprived eye. 163 This effect, however, is only exerted when the eye is closed in infant cats. Reopening the eye at a later time point does not reverse the anatomic effects. Thus, activitydependent plasticity in this area in cats is restricted to a certain period or critical window during development and infancy. Ocular dominance plasticity in the visual system is just one classical example of activitydependent circuit refinement that is temporally restricted to a developmental period; each CNS structure of each species has its own individual critical window. Only within this activitydependent critical window, can the different regions of the nervous system tailor their circuitry to the environment; after closing, circuitry in those regions remains unaffected by changes in activity.
Figure 6 Pum2 is a physiological target of miR-134 during activity-dependent dendritogenesis. (A) Two independent Pum2 shRNAs efﬁciently downregulate the expression of recombinant Pum2. Western blot analysis of HEK293T cell lysates transfected with the indicated amounts of GFP–Pum2 expression vectors, two Pum2 shRNAs as well as a control shRNA (4 ng each) were analysed by western blot using an anti-Pum2 antibody and an anti-eIF4E antibody as a loading control. (B) Si-Pum2-2 efﬁciently downregulates the expression of the endogenous Pum2 protein. Hippocampal neurons were transfected at DIV4 with GFP together with either Si-Pum2-2 (left) or a control siRNA (right, 2 ng each). The cells were ﬁxed at DIV7 and analysed by immunocytochemistry using an anti-Pum2 antibody. Arrows point to cell bodies of representative transfected neurons. Note the reduced Pum2 signal in the cell transfected with Si-Pum2-2 (left) compared with the control neuron (right). Scale bar: 10 mm. (C) Pum2 knockdown by siRNA rescues the miR-134 loss-of-function phenotype in membrane-depolarized hippocampal neurons. Hippocampal neurons were transfected with GFP in conjunction with the indicated anti-miRs (50 nM) and shRNA constructs (2 ng). At 7DIV, neurons were incubated with 16 mM KCl for 6 h and dendritic complexity was evaluated 3 days later. Scale bar: 20 mm. (D) Quantitative analysis of the dendritogenesis assay shown in (C). The calculation of dendritic complexity and the data presentation are as in Figure 4B. Here, 10–16 cells for condition were analysed in each experiment. Data represent the mean of three independent experiments þ s.d. *Po0.05 (t-test). (E) Overexpression of miR-134 duplex RNA perturbs membrane depolarization-induced dendritic outgrowth. Hippocampal neurons (4DIV) were transfected with GFP together with the indicated duplex RNAs (10 nM), and depolarization was performed at 7DIV followed by the assessment of dendritic complexity at 10DIV, as described in (C). Here, 10–16 cells
All of the in vivo studies had in common that behavioral changes of the mice changed the neuronal activity of the relevant sub-region of that behavior and that in turn the neuronal activity then changed the structural spine dynamics. Indeed activity-dependent synaptic strengthening using long-term potentiation (LTP) was shown to induce growth of filopodia in cultured slices of which some were stabilized and maintained ( Engert and Bonhoeffer, 1999; Maletic-Savatic et al., 1999 ). LTP not only showed the induction of newborn spines through filopodia but also influenced the stabilization of new spines ( Lang et al., 2004; Matsuzaki et al., 2004; Nägerl et al., 2004; Hill and Zito, 2013 ). This stabilization and maturation process seems to be linked to the incorporation of PSD-95 into the spine head. The PSD-95 binds both AMPA and NMDA receptors and thereby is an essential factor to convert a nascent spine into a functional synapse. Knockout of PSD-95 resulted in reduced spine stability and immature synaptic function displayed by a reduced AMPAR to NMDAR ratio ( Migaud et al., 1998; Béïque et al., 2006; Ehrlich et al., 2007 ).
Gain-of-function mutations in the tetrodotoxin (TTX) sensitive voltage-gated sodium chan- nel (Nav) Nav1.7 have been identified as a key mechanism underlying chronic pain in inher- ited erythromelalgia. Mutations in TTX resistant channels, such as Nav1.8 or Nav1.9, were recently connected with inherited chronic pain syndromes. Here, we investigated the effects of the p.M650K mutation in Nav1.8 in a 53 year old patient with erythromelalgia by micro- neurography and patch-clamp techniques. Recordings of the patient ’s peripheral nerve fibers showed increased activitydependent slowing (ADS) in CMi and less spontaneous firing compared to a control group of erythromelalgia patients without Nav mutations. To evaluate the impact of the p.M650K mutation on neuronal firing and channel gating, we per- formed current and voltage-clamp recordings on transfected sensory neurons (DRGs) and neuroblastoma cells. The p.M650K mutation shifted steady-state fast inactivation of Nav1.8 to more hyperpolarized potentials and did not significantly alter any other tested gating behaviors. The AP half-width was significantly broader and the stimulated action potential firing rate was reduced for M650K transfected DRGs compared to WT. We discuss the
Another study deciphered the geneti c response to NMDA receptor signaling by whole-genome ex- pression profi ling also using dissociated hippocampal neurons cultured for 10-12 days in vitro. Inducti on of synapti c NMDA receptors using an acti on potenti al bursti ng protocol, inducing long-lasti ng, transcripti on- dependent synapti c plasti city, led to upregulati on of 478 genes. Some of these genes are associated with neuroprotecti on. In contrast mRNA levels of pro-death genes were decreased. A second approach, bath applicati on of glutamate and simultaneous blocking of synapti c NMDA receptors, initi ated cell-death path- ways downstream of extrasynapti c NMDA receptor sti mulati on. This protocol led to acti vati on of transcrip- ti on of 106 genes, only partly overlapping with the genes upregulated in the fi rst assay (Zhang et al., 2007). In vivo genome-wide profi ling identi fi ed about 1000 acti vity-regulated genes in the hippocampus. Genes were classifi ed to fi ve groups based on their expression kineti cs. The genes were identi fi ed by kainic acid induced seizures and scarifi cati on of the mice to obtain hippocampal ti ssue aft er 1, 4, 8 or 24 hours and subsequent microarray analysis. The upregulati on of 24 of those genes was validated by in situ hy- bridizati on (Hermey et al., 2013). One of the validated genes is the GTPase Arl5b which will be described later in more detail and the role of Arl5b in synapti c plasti city is subject of this thesis.
The concept of oxygen- and capacity-limited thermal tolerance has been supported by work on specimens from various marine ectothermic animal phyla from various climate zones (for review see e.g. Pörtner, 2002, 2010, and addressing the Antarctic: Pörtner, 2006). It states, that tolerance to low and high temperatures is set at the highest organisational level, the whole organism. Limiting processes include the capacities of cardiovascular and ventilatory systems to deliver oxygen to the tissues. Within limits these capacities are subject to adaptive adjustments at the molecular, cellular and tissue levels. These should support resting conditions as well as routine rates of activity, growth and reproduction within the temperature range of optimal performance, which defines the biogeographic range of a species (Pörtner and Knust, 2007). Aerobic scope or aerobic functional flexibility, the increment from a rate at rest to an active rate, is thought to be the first process to experience thermal limitation. It may be determined as the increase in metabolic rate during forced activity (metabolic scope for activity, Bennett, 1978; Crear and Forteath, 2000; Fry and Hart, 1948; Rutledge and Pritchard, 1981) or after feeding (specific dynamic action, Robertson et al., 2002), but is also thought to support temperature-dependent growth. Aerobic scope has not been investigated in an array of crustaceans from different latitudes. To date, the hypothesis has mainly been based on investigations of systemic variables at rest in temperate species. In fish the thermal specialization of aerobic scope has also been investigated during exercise (e.g. Farrell et al., 2008). In the face of ocean warming, studies on cold-adapted species have focussed on thresholds of heat tolerance. But to understand the implications of thermal tolerance for biogeography, it is also important to consider the lower temperature thresholds, especially if we want to apprehend the distribution of decapod crustaceans in the Southern Ocean.
different arrival time of sound waves at each ear; the interaural time differences (ITDs) (Goldberg and Brown 1968; Spitzer and Semple 1995; Yin and Chan 1990). Unlike high frequency sound waves, which are always attenuated by the head before being perceived by the contralateral ear, low frequency sound waves do not experience significant sound pressure level changes between the two ears. This statement, however, was demonstrated wrong for very close low-frequency sounds (distance for humans: 1-2m). Within this range, significant near-field ILDs are produced, which can be utilized to localize low frequency sound sources (Shinn-Cunningham et al. 2000). On the other hand, ITDs can also be conveyed by the envelopes of high-frequency sounds and thus elicit similar responses as the fine structure of low frequency sounds alone as observed by in vivo recordings from neurons of the guinea pig IC (Griffin et al. 2005). This study agrees with results reported at the same time indicating that low frequency LSO neurons are sensitive to ITDs (Tollin and Yin 2005). These two findings contradict the strict dichotomy of the “duplex theory” but will be regarded as exceptions for an overall valid hypothesis in this thesis. Hence, generally spoken, sounds composed of wavelengths larger than the head diameter, which is in a first approximation equal to the inter-ear-distance, instead circle around the head before they impinge on the contralateral ear (Fig.1.5). The resulting cue is a well-defined delay dependent on (1) the medium the sound is transmitted in (2) the position of the sound source in the azimuth and (3) the head size. Head sizes of course vary amongst mammals. Larger species exhibit larger ITDs because the time sound waves need to travel from ear to ear increases with increasing inter-ear-distance (Masterton et al. 1967). Humans, for example, experience ITDs of up to 660µs (Feddersen et al. 1957), dogs up to 800µs (Goldberg and Brown 1968; 1969; Grothe and Neuweiler 2000), cats around 300µs (Yin and Chan 1990), whereas some bats, due to their small head size experience maximal ITDs of about 30µs (Pollak 1988).
56 The acute inhibition of PI3K/Akt pathway abrogated the dendritic protrusion formation by Enplastin or Np65 antibody (Np65 Ab). While MG123 inhibition of proteasome activity also prevented dendritic protrusion formation, application of the more specific proteasome inhibitor lactacystin (10 µM) as well as of the protein synthesis blocker or of RelA nuclear translocation did not affect dendritic protrusion formation induced by Enplastin or Np65 Ab (Fig. 29A, B). Collectively, these data indicate that Enplastin- or Np65 Ab-induced early synaptogenesis is dependent on PI3K/Akt but neither on protein synthesis nor the NF-κB pathway. These results also confirm that the extracellular engagement of Np65 is sufficient for its synaptogenic activity in neurons. The aforementioned experiment showed that acute inhibition of RelA did not affect the Enplastin-induced dendritic protrusion formation. Next, I inhibited RelA nuclear translocation chronically for 24 h in neurons. For this, GFP-transfected hippocampal neurons were treated with DMSO or SN50 for 1 day and then Enplastin was applied for 1 h. Interestingly, long-term inhibition of RelA did not affect the Enplastin-mediated increase in dendritic protrusion numbers in neurons (Fig. 30A, B), suggesting that this process is independent of RelA signaling in young neurons.
Substituted phenylpyridazinones can act as “bleaching” herbicides, i.e., they interfere with ca rotenoid biosynthesis and prevent the subsequent build-up of chlorophyll and other constituents of the thylakoid . This phytotoxic effect depends on various substituents [2 -4 , see the latter for review]. In a recent publication, bleaching activity of such com pounds was quantitatively assayed by measuring their interference with the greening process, using a sensitive Scenedesmus m utant . As pointed out previously , this algal model system allows for fast and reliable data on structure-activity relation ships. We found that the prom inent effect of the substituents is by influencing the charge distribu tion of the phenylpyridazinone molecule. Pigm ent bleaching improved firstly by substituents with increasing am values at position 4, secondly with decreasing ov values at position 5 of the pyrida- zinone ring, and thirdly by substituents with in creasing am or Up values at the m eta position o f the phenyl moiety (; see formula in Table).
If the Paranormal Activity films are “accelerationist”—or if an accelerationist aesthetic is at work in post-cinematic production more generally—then this is because, in order to present us with impalpable demonic forces, these recent films are compelled to adopt, and adapt to, the most cutting-edge tendencies of actually existing capitalism. It has often been suggested that classical Hollywood continuity editing instantiates the same logic as Fordist-Taylorist industrial mass production. I think that, similarly, the editing practice of contemporary film and video production instantiates the same logic as does the post-Fordist regime of flexible, just-in-time production (best described by David Harvey). Under this regime, David Bordwell’s “intensified continuity” has hyperbolically extended itself, and thereby mutated, into what I have called in my book “post-continuity.” The classical norms of smooth narrative development and intelligible scene construction are no longer in force. Indeed, throughout contemporary film production, these norms are violated in opposite directions at once. In the post-cinematic, we find both excessive movement (shaky cameras), and excessive stillness (fixed cameras). We find both baroque narrative elaboration and complexification, and the abandonment of narrative or causal logic altogether. And we find both an exceedingly literalistic psychology of character development, in which every last tic and affectation must be given a “plausible” motivation, and the complete abandonment of any sort of character development or motivation whatsoever. Both extremes are affirmed at the expense of the classical norm or mean.
822 S. M üller and A . Schm idt • Su bstrate-D ependent Arylsulfatase A ctivity
sulfatase activity during sulfur sta rv a tio n co n d itio n s, in c o n tra st to tw o green algal strains. H o w ev er, sig nificant arylsulfatase activity was d ete c te d during grow th on ethan esu lfo n ic acid and th e sulfate e ste r am in o eth y l sulfate. B est yields w ere o b ta in e d using th e su b stra te p -n itro p h en y l su lfate, w hereas no activ ity was d e te c te d using th e co rresp o n d in g p -n itro - catech o l sulfate as sulfur source fo r grow th. It is evi d e n t th a t no arylsulfatase could be d etec ted during “n o rm a l” sulfur starvation o r on any sulfur co m p o u n d in th e presence of sulfate. T hus a sulfur s ta r vatio n signal and th e presence o f th e c o rrec t su b stra te (in d u ce r? ) are necessary fo r o p tim al ary lsu l fata se d ev e lo p m e n t. A lgae could be grow n e ith e r on n o rm al air o r using air en rich ed w ith 5% C 0 2 w ith o u t alterin g th e results concerning arylsulfatase activ ity (d a ta n o t show n).
particularly sensitive to the local-type primordial non-Gaussianity. Specifically, we divide a survey into subvolumes, and measure the correlation between the position-dependent two-point statistics and the long-wavelength overdensity. This correlation measures an in- tegral on the bispectrum, and is dominated by the squeezed-limit signal if the wavenumber of the position-dependent two-point statistics is much larger than the wavenumber cor- responding to the size of the subvolumes. Therefore, without employing the three-point function estimator, we can extract the squeezed-limit bispectrum by the position-dependent two-point statistics technique. Furthermore, nonlinearity of the correlation between the position-dependent two-point statistics and a long-wavelength mode can be well modeled by the separate universe approach, in which the long-wavelength overdensity is absorbed into the background cosmology; the window function effect can also be well taken care of because this technique measures essentially the two-point function and the mean overden- sity, for which the procedures for removing the window function effects are relatively well developed. With the above advantages, the position-dependent two-point statistics is thus a novel and promising method to study the squeezed-limit bispectrum of the large-scale structure.
original mudpot water containing particular high amounts of dissolved lanthanides or in medium supplemented with lanthanide salts.  SolV expresses the same xoxF encoded MDH as mentioned above, metabolizing methanol for energy generation and containing a lanthanide ion in its active site.  Several clades of XoxF have evolved in different microorganisms, commonly named XoxF1-5. [37-39] The name is derived from the homology to calcium dependent MxaF-type MDH and the x stands for the unknown function at that time.  xoxF is positioned on an operon, together with xoxG, encoding the natural electron acceptor for MDH and xoxJ, encoding a binding protein of unknown function.  The crystal structure of the enzyme isolated by Pol et al. was refined with cerium in the active site, although the metal ion in this enzyme was a Ln mixture due to mudpot water used. The central metal is coordinated by amino acids Glu172, Asn256, Asp299 and Asp301 and the known cofactor pyrroloquinoline quinone (PQQ), an organic heterocyclic molecule, functionalized with three carboxyl groups. 
Overall, these results suggest that the safener fenclorim may enhance the metabolic detoxication o f pretilachlor via conjugation to glutathione by stim ulating the direct or indirect synthesis o f gluta thione rath er than the activity of the GST enzyme catalyzing this reaction. In addition, these results suggest th at a sim ultaneous application of preti lachlor and fenclorim is critical for the enhance ment o f glutathione and glutathione-related en
Fig. 2. T em perature stability of C su b u n it o f cA -protein kinase II from rat muscle in absence (A), and in presence (B) of substrate histone or cosubstrate ATP. C subunit preparations diluted properly with a 0.1% (w/v) solution o f bovine serum album in were incubated in siliconized glass tubes: (A) at 26 °C ( • ) , 30 °C (O), 35 °C (A), 40 °C (□ ), 50 °C (■), or 60 °C (A ) in a water bath under gentle shaking. Samples were w ithdrawn after 0 (control), 1, 2, 3, 5, 7, and 20 m in and placed on ice (Eppendorf reaction tubes). (B) at 40 °C either alone (O) or in presence of histone (□ ), o f A TP in absence o f Mg2+ (A ), or o f A TP in presence o f M g2+ (V ). Ratios o f mixtures as well as concentration o f M g2+ were as in the standard assay. This test was carried o u t identically at 50 °C: enzyme alone ( • ) , m ixtures of enzyme and histone (■ ), o f enzyme and ATP in absence (A) or presence (▼) o f M g2+. Samples were withdrawn after 0 (control), 1, 5, and 10 m in and placed on ice (E ppendorf reaction tubes). Kinase activity was assayed in triplicate according to the standard procedure using histone as substrate. Activities found were related to the respective control. To test for the stability o f substrate and cosubstrate, histone in absence ( - x - ) and presence (— x — ) o f 0.1% bovine serum album in, and ATP in absence (—I — ) and presence (------ 1 -------) o f 10 mM M g2+ were incubated at 50 °C and em ployed in
The issue is that no American, Japanese, or European cities have experienced a motorcycle dependent situation. Therefore, the consultants carried out their works mainly by the car- based planning methods, tools, and experience. The planning solutions were normally focused on road development, except few comprehensive transport planning studies, for example the Bangkok Transport Study 1975 (Rujopakarn, 1999). The foreign defined objectives are far from implementation abilities and political interests of local implementers. The urban road network was planned and designed for the car traffic only, while the car- based transport system cannot operate safely and efficiently with a mixed traffic flow. Simple conversion from motorcycle or bicycle to passenger car unit has many limitations. Based on the passenger car unit and level of service for car performance, the planned road capacity in the future is usually much larger than the actual need. If the plan were realised, the city would have to spend a huge amount of money, mainly from foreign loans, and large area of land for that expressway, which would depend much more on car use. The social and environmental impacts will also be a very big problem. If the plan is not implemented, the city would have no preparation for meeting the future traffic demand Therefore, the current transport problems would remain, and the new problems would be generated.
In this chapter the basic concepts necessary to understand spin-dependent transport phenomena in magnetic nanocontacts are introduced. First sec- tion (Sec. 2.1) deals with magnetic domains and domain walls (DWs). Two types of domain walls are described: bulk and geometrically confined DWs. The latter appears in magnetic constrictions and represents a new kind of DW, besides the unconstrained Bloch and N´eel bulk walls. These systems are characterized by means of magnetoresistive effects, which serve as an indirect sensing tool on the nanometer scale. Anisotropic magnetoresis- tance (AMR) caused by spin-orbit interaction and domain wall magnetore- sistance (DWMR) due to scattering of electrons at the domain walls are used to observe the magnetization reversal processes in cobalt structures with constrictions. The AMR effect is presented in Sec. 2.2, followed by a detailed description of the magnetic reversal for a µm large ferromagnetic sample in Sec. 2.3. Then, the correlation between AMR and magnetization reversal is discussed on the basis of an experimental work taken from litera- ture. The theoretical models used to explain the DWMR effect are given in Sec. 2.4. There are two regimes of DWMR defined as function of the DW width, namely, the diffusive regime applicable for nm wide nanocontacts and the ballistic DWMR in the case of atomic point contacts. The diffusive DWMR is discussed to a larger extent, because it is relevant for the exper- iments presented in this thesis. The ballistic DWMR is described in order to give the reader a more complete picture of the spin-dependent transport in nanocontacts down to the atomic scale. An overview of the experimental results obtained on nanocontacts by other groups, in both ballistic and diffu- sive regimes, is given in Sec. 2.5. In the last section (Sec. 2.6), a description of the important issues concerning micromagnetic calculations is presented. The simulations were used to get a better understanding of magnetization reversal processes and their influence on the spin transport characteristics.