In light of the lack of overt evidence proving that the mammalian organ of Corti is able to form new sensory cells, and given the fact that these cells are few and may be formed only once in a life span the significance of the research lies in understanding the mechanism of survival and recovery of receptor cells after injury. Specifically, we propose to study: 1 Post-traumatic cell proliferation in the organ of Corti: A Mitotic cell proliferation in the injured cultures will be examined using 3H thymidine to investigate the identity of dividing cells, the duration of the mitotic response and the age- related occurrence of cell division; B Nonmitotic cell proliferation, expressed by extensive sprouting of supporting cells, will be investigated in search of possible diffusable growth factors NGF, EGF, FGF released by the cells in response to direct injury or to the injury of sensory cells.
The study will encompass the morphological sequence of the regenerative phenomena, its comparison with normal development, and the influence of age on the regenerative capacity of sensory cells. The study will be done using short- and long-term cultures of the mouse organ of Corti. The injury will be done by hand, using either a pulled- glass pipettes or a laser. Light and electron microscopy, autoradiography, biochemistry and immunocytochemistry will be employed.
Some specialized immunological assays will be done in collaboration with Dr. Philippe Lefebvre, University de Liege, Belgium. Toggle navigation.
Principles of Regenerative Biology
Recent in Grantomics:. Recently viewed grants:. Figure 8. Currents, EDR, pressure, voltage versus time in session The time evolution of the total voltage between the electrodes is shown in the last plot of figure 8. The microdischarge onset occurred when the voltage of the negative power supply was decreased from kV to kV. At constant voltage, micro-discharges tended, as usual, to extinguish until a further step-down voltage was applied. These series of micro-discharges were accompanied by the local heating of the anodic support as observed in the previous experimental session see figure 7.
The dynamic response of the x-ray counts detected by the PVT scintillators is in agreement with the other measured quantities but the interpretation of the energy associated to each count is not straightforward. The energy recorded is mainly the result of the Compton scattering [ 11 ] interaction of the primary photons coming from the experiment and the scintillator material. The energy of the primary photons is due to the Bremsstrahlung radiation of electrons decelerated on anode.
 The Regenerative Vibrations Influence on the Mechanical Actions in Turning
Their energy distribution can be described by the Kramer's law [ 12 ]. This diagnostic system and the measurements interpretation will be discussed in detail in [ 9 , 10 ]. Figure 9. Currents, voltages, x-ray spectrum, pressure, EDR as function of time. Figure X-ray spectrums in time windows W 1 , W 2 and W 3 as shown in figure After several tests were executed changing the gap length and the electrode geometry, while the power supply polarity kept unaltered, a couple of annular brown traces have been observed on the vacuum chamber inner wall, in front of the anodic support see figure Inner wall of the HVPTF vacuum vessel: two circumferential stains have been progressively appeared during the execution of the high voltage tests.
A reduced amount of material has been also deposited in the insulator indicated in the point P of figure In figure 13 the sample and the analyses results are shown. Fe and Cr are fully consistent with the composition of the stainless steel which is the sole metal adopted for the vacuum chamber and for the electrode under test. The most obvious conclusion is, thus, that the brown traces are produced by material coming from the stainless steel anodic support. The most evident aspect, concerning the results so far reported in the previous section, is that a spurious phenomenon has influenced the high voltage tests.
The microdischarge activity has involved mainly the vacuum vessel and the positive power supply rather than the two electrodes shown in figure 2 , although they were separated by the shortest gap length and subjected to the largest difference of electric potential. Adopting this hypothesis, it is easy to demonstrate that breakdown should occur when the following relation is satisfied. Direct measurements of coefficients A and B carried out by the same authors [ 13 ] and moreover by [ 14 ] would not immediately support the validity of 1.
Nevertheless, the same type of analysis should be carried out also for the coefficients C and D so as to have a complete benchmark of this assumption with the reality. The energy spectra shown figure 10 has been measured using a plastic scintillator material, so photons interact by single Compton scattering rather than photo-absorption. This means that full energy peaks are not directly observable. Moreover, it is plausible that the distribution of photons generated by the Bremsstrahlung radiation of mono-energetic electrons impinging the anodic surface follows the Kramer law.
Assuming the Compton scattering distribution according with the Klein—Nishina equation [ 15 ], it is possible to reconstruct, by means of a Monte Carlo 1D algorithm, the shape of the spectra shown in figure 10 ,. The numerical results are reported in figure The results of the simulations are consistent with the general trend of the experimental results, which exhibits an energetic spectrum having negative slopes with a maximum energy according with the accelerating voltage.
Results of the Monte Carlo calculation: reconstruction of the x-ray spectra recorded by the EJ scintillator due to the bremsstrahlung radiation of a monoenergetic electron beam of keV impinging the vacuum vessel. The number of the counts of the simulations is proportional to a coefficient which could be obtained by specific tests accelerating a known electron current toward the anode and measuring the rate of the x-ray counts detected by the scintillator. In this view, the microdicharge activity would lead toward a progressive reduction of the electron extraction from the cathodic surface, probably due to not negligible contribute of the extracted ions from the anodic surface.
The peculiar spatial distribution of the emission sites on the electrode surfaces, shown in figure 7 , has been analyzed thanks to the idea proposed in [ 16 ] where a mutual exchange of charged particles, with opposite sign, is simulated between electrodes having a generic shape. The numerical simulations reported in [ 16 ] reveal the existence of specific points on the electrode surfaces able to concentrate the charged particle exchanged between electrodes. A set of numerical simulations considering the HVPTF geometry has been carried out, following the same approach, to justify the observations described in the previous section.
The trajectories of positive charged particles, uniformly distributed on the anodic surfaces, have been integrated considering the 2Daxialsimmetric electrostatic field map reported in figure 3. The axial-symmetric domain assures the conservation of the angular momentum. The starting positions of the charged particles have been located on the anode surface on the right hand side.
The integration of the equation of the motion starts assuming an initial null speed: this assumption assures trajectories having a radial path located in a plane at constant azimuthal coordinate. The trajectory stops when a charged particle hit an electrode, than the sign of the charge is changed and a new trajectory is integrated.
The process has been repeated up to a condition characterized by a stationary mutual-exchange of trajectories. In this geometry, twenty iterations are necessary to converge toward the final configuration. The trajectories have been grouped and exchanged only between two points A and C, respectively on the anode support and the cathodic surface of the vacuum vessel.
Such points are called discharge attractors as referred in [ 16 ]. It is possible to demonstrate that the trajectory path of a charged particle in an electrostatic irrotational field, assuming classic motion and zero initial velocity, depends only on the shape of the domain and the ratio of the applied voltages in case of multi-electrode system.
Application of biofabrication technologies
A concise demonstration of the independence of the trajectory upon mass, charge and even gap voltage is reported in Appendix. Under such conditions the attractor locations depend only on the shape of the electrodes and on the ratio between the applied voltages. The first three and the last 20th iterations are shown in figure 15 : the trajectories of the positive charged particles are in red, while the ones of the negative charges are in black. The numerical analyses above described adopt a very simple model, based on restrictive hypotheses of a null initial velocity.
Nevertheless, dedicated high voltage tests with double polarity could be designed in order to discern this important aspect.
The analyses of experimental results in the following situation: and or vice versa should be sufficient to understand the asymmetric behavior of cathode versus anode during the high voltage conditioning in vacuum. The electrode configuration discussed so far has symmetric domain boundaries except for the region between the electrodes under test, thus another couple of attractor points exists on the support on the left.
However, as anticipated in the previous section, no detectable temperature rise has been observed on the structure connected to the negative power supply neither on the vacuum chamber wall. This is confirmed by the generally small current I- recorded during the experimental sessions. This asymmetric behavior depends on the different role assumed by anodic and cathodic surfaces in the micro-discharges onset.
In spite of many investigations carried out in the past, a consolidated model describing the physical phenomena occurring during the high voltage conditioning over long vacuum gaps is still missing in literature.
Nevertheless the analyses of the present experimental results would suggest that the micro-discharges activity would depend on an cathode-anode interaction in which the local heating of the anode play a fundamental role in the micro-discharge onset. Probably the large anodic surface in front of the support connected to the negative power supply on the left is sufficient to limit the local temperature and the local outgassing. The same does not occur on the cantilever support connected to the positive power supply.
Actually the RGA plot of figure 6 is compatible with the measurements reported in [ 17 ]. The same type of emissions could be observed if the gas desorption had been thermally activated, nevertheless specific tests to confirm this hypothesis have not been done yet in our high voltage test stand.
The electrode shape could quickly focus the exchanged particles toward the accumulation points. Even though the latter generally are not located in the highest electric field area, the accumulation points can nevertheless collect the contribute of a large portion of electrode. The high flux of heat can rise locally the temperature enhancing the electrode sputtering, the desorption of the gases trapped in the layer on the metal vacuum interface and eventually promoting the microdischarge onset. Sketch representing the cascade particles exchange between two generic electrodes, the red paths are the trajectories of the positive ions while the black one are the trajectories of the negative charged particles a layer of adsorbed gases cover the surface of both electrodes.
The mechanism so far described is one of the possible processes which limit the voltage holding of a generic system insulated by vacuum gaps. Experimental evidences concerning the existence of accumulation points during occurrence of micro-discharges have been observed during the high voltage conditioning of an electrostatic device insulated by large vacuum gaps. A mutual exchange of positive-negative charged particle is the mechanism which presumably causes this phenomenon and defines the position of the micro-discharge attractors.
The attractors regions due to the regenerative processes of charged particles between cathode and anode have been identified thanks to numerical ray-tracing simulations. The position of the attractors depends only on electrode shape and the ratios of applied voltages. A perturbation of the trajectories position by altering the voltage distribution has been sufficient to initiate the micro discharge occurrence.
The existence of phenomena able to concentrate the dissipation of power in specific location on the electrode surfaces during the high voltage conditioning in vacuum could be exploited in the development of neutrons sources applications. The particles exchange could involve only electrons and positive ions or might be supported also by the formation of negative ions as postulated in [ 16 ].
In this second hypothesis, the phenomena would imply also the migration and the accumulation of material on the discharge attractors. Thus, the formation of accumulation points could be exploited in the development of neutrons sources applications. Attempts to generate neutron sources by electrostatic accelerators by means of the bombardment of fast deuterium ions to hydrogen isotopes adsorbed in solid targets such as titanium have already been carried out for decades.
The following sentence has been extracted from a '60 paper describing the development of a sealed-off vacuum tube used as sources of neutrons from fusion reaction [ 18 ]. It is interesting to observe that the attempt to generate positive ions was limited by the high voltage conditioning of the hydrogenated titanium electrodes adopted in such specific test. Considering the observations reported in the present paper, a further step forward in the development of neutrons sources applications could be the adoption of optimized system of electrodes to enhance the formation of attractor points.
The accumulation of charged ions toward specific points could improve the efficiency of the neutron source limiting the loss of reactants sputtered by the accelerated ion collision. The mechanism could be effective if the production of negative hydrogen ions on the cathodic surface is larger than the extraction of electrons, this could be obtained by combining two effects: seeding cesium at the cathode side and applying magnetic filter field for electrons in a region enclosing the attractor on the cathodic surface.
The proposal here reported is a speculation which can be verified only by dedicated experimental campaign. The independence of the classic trajectory upon mass, charge and even gap voltage can be done by means of the Buckingham Theorem [ 19 ]. Referring to figure A1 , the distance L the variable identifying the ending point of the trajectory is a function of all the physical parameters governing the system. Figure A1. Quantities determining the throw L of a charged particle in an electrostatic field map, example of sphere plane geometry. The number of fundamental physical units involved in equation 3 is 4, it corresponds to: [A],[s],[kg],[m] while the number of implicit variables in the same equation is 7.
Author e-mails. Peer review information. In case of plane parallel electrode the maximum voltage holding is proportional to the gap length and thus The latter equation is not valid in the long gap cases where with [ 5 ] [ 6 ]. Zoom In Zoom Out Reset image size. Figure 4. Automatic conditioning procedure parameters.