The effect of polypyrrole with incorporated neurotrophin-3 on the promotion of neurite outgrowth from auditory neurons

This research aims to improve the nerve-electrode interface of the cochlear implant using polymer technology to encourage neuron survival, elongation and adhesion to the electrodes. Polypyrrole (Ppy) doped with p-toluene sulphonate (pTS) is an electroactive polymer into which neurotrophin-3 (NT3) can be incorporated. Ppy/pTS+/-NT3 was synthesised over gold electrodes and used as a surface for auditory neuron explant culture. Neurite outgrowth from explants grown on Ppy/pTS was equivalent to tissue culture plastic but improved with the incorporation of NT3 (Ppy/pTS/NT3). Electrical stimulation of Ppy/pTS/NT3 with a biphasic current pulse, as used in cochlear implants, significantly improved neurite outgrowth from explants. Using (125)I-NT3, it was shown that low levels of NT3 passively diffused from Ppy/pTS/NT3 during normal incubation and that electrical stimulation enhanced the release of biologically active NT3 in quantities adequate for neuron survival. Furthermore, Ppy/pTS/NT3 and its constituents were not toxic to auditory neurons and the Ppy/pTS/NT3 coating on gold electrodes did not alter impedance. If applied to the cochlear implant, Ppy/pTS/NT3 will provide a biocompatible, low-impedance substrate for storage and release of NT3 to help protect auditory neurons from degradation after sensorineural hearing loss and encourage neurite outgrowth towards the electrodes.     

Graft-polymerization of styrene on polyethylen single crystals

The graft polymerization of styrene through use of γ-ray irradiation was carried out in vapour phase onto polyethylene single crystals, bulk films of polyethylene and paraffine crystals. It was found that the grafting yield for a given dosage was much less for polyethylene single crystals than for bulk films. The similar difference was also found between the pure paraffine (n-tetracosane) and mixed paraffines, and these results were interpreted in the light of crystallinity. In order to characterize the polystyrene branches grafted onto the crystals, a new method based on chain transfer reaction was proposed for obtaining number of the grafted points and the length of the branches and was applied to the graft polymerization onto the polyethylene single crystals. The results were compared with those obtained for the graft polymerization onto paraffine crystals. In the latter case, molecular weight of the graft polymer obtained after extracting paraffines from the whole material could be obtained by assuming the copolymer as polystyrene homopolymer. In both of the cases the polymer branch was very long and the sites of grafting were very few. The crystallinities of the samples measured by a X-ray diffraction method before and after the grafting showed that the grafting did not practically affect the original crystals. The surfaces of the grafted crystals were also examined by an electron microscope and the result implied that the reaction took place mainly on the surfaces of the crystals.     

Crystallization of polypeptides in the course of polymerization, 6. Effect of the heteroatom attached to the β-carbon on the crystal growth

In order to examine the growth mechanisms of polypeptides with a heteroatom, S, on the β- and γ-carbons which give the α-conformation and the α-helical conformation, resp., in the course of a heterogeneous polymerization, we have studied kinetics of the heterogeneous polymerization of S-methyl-L-cysteine N-carboxy anhydride (NCA), S-benzyloxycarbonyl-L-cysteine NCA and L-methionine NCA initiated by butylamine in acetonitrile and the morphologies of the resultant polymers. As was anticipated, it has been verified that poly-(S-substituted-L-cysteines), which take only the β-structure as the ordered form, gave the chain-folded crystals. In all the cases studied, long lath-like crystals formed in the early stages of polymerization thickened into round rod-like fibers as the polymerization proceeded. A possible model was proposed for the growth mechanism of these polymer crystals, taking into account the cross-β-structure. It has been revealed that poly(L-methionine) with the α-helical structure gave the extended chain crystals.     

Kinetik und mechanismus der kationischen polymerisation von trioxan: Ein katalysierter kristallwachstumsprozeß

The kinetics of the cationic polymerization of trioxane initiated by perchloric acid is analyzed as a simultaneous polymerization and crystallization process. The details of the crystal growth such as nucleation, distribution of crystal sizes, development of growth spirals etc. have been investigated by electron and light microscopic techniques. The mechanism of polymerization can be understood as a catalyzed crystal growth. A well defined population of crystals is formed during a nucleation event at the start of the polymerization. An adsorption equilibrium with regard to the catalyst is then established in the presence of the rather large surface area of the growing polymer crystals. Assuming a Langmuir-type adsorption behaviour, the observed growth features (growth spirals) can be quantitatively interpreted on the basis of the BCF-theory of crystal growth. The current concepts on the molecular mechanism of cationic polymerization of cyclic formals have been modified such as to take into account that each chemical step of the polymerization is a building step of the crystal too.     

Anionic polymerization of siloxanes. Mechanism of initiation with triorganosilanolates

The kinetics of initiation of the polymerization of 2,2,5,5-tetramethyl-1-oxa-2,5-disilacyclopentane ( 2 ) with sodium phenyldimethylsilanolate 1a and two of its phenyl substituted derivatives was investigated as model reaction for studying the initiation and crosspropagation steps in anionic polymerization of cyclic siloxanes in non-polar media. A redistribution of siloxane units between living ends of the polymer and also between living end and initiator was demonstrated to occur commonly in the siloxane anionic polymerization system affecting the initiation and crosspropagation. However, the living silethylenesiloxane polymer system 3 showed no redistribution and the initiation, followed by the dissapearance of phenyldimethylsilanolate, showed a first internal order and a fractional external order with respect to the initiator. The same fractional order was found in the propagation step with respect to the living ends. The results can be rationalized on the basis of a mechanism in which silanolate groups aggregation (cyclic or cage structures) convert active species into dormant centres. The analysis of the kinetic scheme including the formation of simple and mixed aggregate series of optional multiplicity leads to the conclusion that the scheme under some assumptions suits the kinetics observed. Solvation processes as well as the increase of the negative charge density on the silanolate oxygen by electron releasing substituents increase the reactivity of the silanolate.     

The solid-state polymerization,physical properties,and crystal structures of diacetylene mixed crystals

Suitable substituted diacetylenes, 2,4-hexadiynylene di-p-toluenesulfonate (1a) and 2,4-hexadiynylene di-p-fluorobenzenesulfonate (1b) , were cocrystallized to form substitutional solid solutions. By solid state polymerization of mixed crystals of 1a and 1b single crystals of the corresponding copolymers were obtained. The polymerization rate can be varied over a wide range by admixture of the comonomer 1b in the lattice of 1a . The crystal structures of monomer 1b and of the corresponding polymer, 2b , were determined. It was found that the packing of the reactive triple bond systems is identical in both monomers. Comonomer 1b acts as a defect site with a high probability of chain termination.     

Polypyrrole-coated electrodes for the delivery of charge and neurotrophins to cochlear neurons

Sensorineural hearing loss is associated with gradual degeneration of spiral ganglion neurons (SGNs), compromising hearing outcomes with cochlear implant use. Combination of neurotrophin delivery to the cochlea and electrical stimulation from a cochlear implant protects SGNs, prompting research into neurotrophin-eluting polymer electrode coatings. The electrically conducting polypyrrole/para-toluene sulfonate containing neurotrophin-3 (Ppy/pTS/NT3) was applied to 1.7 mm² cochlear implant electrodes. Ppy/pTS/NT3-coated electrode arrays stored 2 ng NT3 and released 0.1 ng/day with electrical stimulation. Guinea pigs were implanted with Ppy/pTS or Ppy/pTS/NT3 electrode arrays two weeks after deafening via aminoglycosides. The electrodes of a subgroup of these guinea pigs were electrically stimulated for 8 h/day for 2 weeks. There was a loss of SGNs in the implanted cochleae of guinea pigs with Ppy/pTS-coated electrodes indicative of electrode insertion damage. However, guinea pigs implanted with electrically stimulated Ppy/pTS/NT3-coated electrodes had lower electrically-evoked auditory brainstem response thresholds and greater SGN densities in implanted cochleae compared to non-implanted cochleae and compared to animals implanted with Ppy/pTS-coated electrodes (p < 0.05). Ppy/pTS/NT3 did not exacerbate fibrous tissue formation and did not affect electrode impedance. Drug-eluting conducting polymer coatings on cochlear implant electrodes present a clinically viable method to promote preservation of SGNs without adversely affecting the function of the cochlear implant.     

Inverse microemulsion polymerization of acrylamide in the presence of hexamethylenetetramine

The inverse microemulsion polymerization of acrylamide is retarded in the presence of hexamethylenetetramine. The retardation effect of a given hexamethylenetetramine concentration on the kinetics of the polymerization of acrylamide depends on the pH of compartmentalized water droplets. For non-crosslinked polymer particles the polymer particle diameter for a given composition of the inverse microemulsion increases in the presence of hexamethylenetetramine, while the relative molecular mass (molecular weight) of the polymer decreases. Depending on the pH value of the compartmentalized water droplets polymer particles with different crosslinking degree (of different swelling index) were obtained. The kinetics of polymerization and the polymer particle characteristics are discussed with respect to the hexamethylenetetramine effect on the stability of dispersed particles in an inverse microemulsion prior to and during the polymerization and in terms of the effect of “in situ” formed hexamethylenetetramine decomposition products and of their reactions with acrylamide and/or acrylamide structural units in the polymer.     

Polymerization of acrylamide photoinitiated by cobalt ammine complexes

The kinetics of polymerization photoinitiated by pentaammineoxalatocobalt(III) and tetraamminebis(oxalato)cobalt(III)complexes were investigated in aqueous and mixed solvents by irradiation of these complexes in the charge-transfer bands at 35°C. Radiation of wavelength 420 nm was employed and the kinetics were followed by measurement of the rate of monomer disappearance (gravimetrically) and chain lengths of polymer formed (viscometrically). The dependence of the rate of polymerization and of the chain lengths on factors like absorbed light intensity, light absorption fraction, monomer concentration, initiator concentration, hydrogen ion concentration, viscosity of the medium, and the nature of the outer anion were studied in detail. A kinetic scheme is proposed and discussed in the light of experimental results, involving (i) a primary photo-chemical act of excitation of the complex, followed by dark reaction of electron transfer within the complex, producing free radicals and bivalent Co²⁺ ions, (ii) initiation of polymerization by the free radicals generated, and (iii) termination of the active chain by a mixed process, namely mutual termination of macroradicals as well as termination by the complex depending on experimental condition.     

Solid state polymerization of a nitrophenoxy disubstituted diacetylene

The solid state polymerization of the disubstituted diacetylene 1,6-bis(2,4-dinitrophenoxy)-2,4-hexadiyne by thermal annealing at 130°C to yield high quality polymer single crystals is reported. This compound is unique in that it polymerizes by thermal annealing but not under ordinary x-ray conditions, thus allowing determination of the crystal structures of both the monomer and polymer by x-ray diffraction. The conditions for solid state polymerization and the structural changes during polymerization were investigated. In addition, the kinetics of polymerization were studied by differential scanning calorimetry. An induction period with activation energy E_A(slow) = 139,7 ± 3 kJ/mol (33,4 ± 0,7 kcal/mol) is followed by a rapid polymerization regime with activation energy E_A(fast) = 132,2 ± 6 kJ/mol (31,6 ± 1,5 kcal/mol) during which first order kinetics are obeyed. The total heat of polymerization is ? ΔH_p = 134,7 ± 6 kJ/mol (32,2 ± 1,5 kcal/mol). After polymerization, a disordering phase change takes place with continued isothermal annealing which becomes clearly evident at isothermal annealing temperatures higher than 130°C.     

The solid-state polymerization and physical properties of 2,4-hexadiynylene bis(N-ethylcarbamate), 1 crystal modifications and polymerization kinetics

Three distinct crystal modifications of 2,4-hexadiynylene bis(N-ethylcarbamate), ( 1c ) are isolated under different conditions. The conditions necessary for the growth of crystals of the different modifications are described and their melting and phase transitions determined by differential thermal analysis. X-Ray diffraction studies yielded unit cell dimensions for some modifications of the monomer 1c as well as the polymer 2c , but full structural analyses were impossible to obtain. Molecular models of 1c with different conformations have been constructed and shown to be compatible with the observed cell dimensions and solid-state reactivities. The thermal and radiation polymerization kinetics of the two reactive modifications of 1c were studied. Despite the fact that the thermal polymerization of modification I of 1c is not a single-phase process, the kinetics are well described by the crystal-strain model, this fit being clearly fortuitous. A more detailed study of the kinetics reveals a second-order behaviour and a model for a two-phase polymerization process is proposed.     

Kinetische und morphologische untersuchungen zur polymerisation von oxacyclischen monomeren im festen zustand, 5. Zur morphologie der während der sublimation von trioxan- und tetroxankristallen wachsenden polyoxymethylene

The polymerization and crystallization of polyoxymethylene obtained by sublimation of trioxane (1,3,5-trioxane) and tetroxane (1,3,5,7-tetroxocane) crystals was investigated. Electron microscopy techniques and thermal analysis were used to characterize the polymer crystals as produced and after annealing. The polyoxymethylene crystals resulting from tetroxane show a regular lamellar folded chain macroconformation similar to those previously reported by polymerization of tetroxane crystals chemically initiated. The polymerization follows a crystallization succeeding polymerization mechanism. Trioxane gives mainly fibrous crystals, showing well oriented electron diffraction diagrams and higher melting points, consisting mostly of extended chains. A “pseudolamellar” morphology was also observed, being established by annealing, that the chain directions are parallel to the edges of the crystals. The strange growing habit seems to be produced by disruption of the electrical field of the trioxane crystals by a water effect. The polymerization seems to follow mainly a simultaneous polymerization and crystallization mechanism. The initiation of the polymerization is in both cases due to the gaseous formaldehyde formed during sublimation.     

Promoting neurite outgrowth from spiral ganglion neuron explants using polypyrrole/BDNF-coated electrodes

Release of neurotrophin-3 (NT3) and brain-derived neurotrophic factor (BDNF) from hair cells in the cochlea is essential for the survival of spiral ganglion neurons (SGNs). Loss of hair cells associated with a sensorineural hearing loss therefore results in degeneration of SGNs, potentially reducing the performance of a cochlear implant. Exogenous replacement of either or both neurotrophins protects SGNs from degeneration after deafness. We previously incorporated NT3 into the conducting polymer polypyrrole (Ppy) synthesized with para-toluene sulfonate (pTS) to investigate whether Ppy/pTS/NT3-coated cochlear implant electrodes could provide both neurotrophic support and electrical stimulation for SGNs. Enhanced and controlled release of NT3 was achieved when Ppy/pTS/NT3-coated electrodes were subjected to electrical stimulation. Here we describe the release dynamics and biological properties of Ppy/pTS with incorporated BDNF. Release studies demonstrated slow passive diffusion of BDNF from Ppy/pTS/BDNF, with electrical stimulation significantly enhancing BDNF release over 7 days. A 3-day SGN explant assay found that neurite outgrowth from explants was 12.3-fold greater when polymers contained BDNF (p < 0.001), although electrical stimulation did not increase neurite outgrowth further. The versatility of Ppy to store and release neurotrophins, conduct electrical charge, and act as a substrate for nerve-electrode interactions is discussed for specialized applications such as cochlear implants.     

Impact of co-incorporating laminin peptide dopants and neurotrophic growth factors on conducting polymer properties

Conductive neural interfaces tailored for cell interaction by incorporation of bioactive factors are hypothesized to produce superior neuroprostheses with improved charge transfer capabilities. This study examined the effect of entrapping nerve growth factor (NGF) within the conducting polymer poly(ethylene dioxythiophene) (PEDOT) during electrodeposition to create a polymer capable of stimulating neurite outgrowth from proximal neural tissue. NGF entrapment was performed on polymers doped with laminin peptides DEDEDYFQRYLI and DCDPGYIGSR and, additionally, a conventional dopant, paratoluene sulphonate (pTS). All polymer coatings were analysed for a range of physical, electrical and mechanical properties, with the biological activity of ligands examined using a PC12 neurite outgrowth assay. NGF was successfully entrapped in PEDOT during electrodeposition and was shown to produce a softer interface than conventional conducting polymers and films without the NGF modification. However, it was found that the use of a peptide dopant combined with NGF entrapment resulted in polymers with diminished electrical and mechanical stability. Entrapped NGF was determined to be biologically active, with PEDOT/pTS/NGF producing neurite outgrowth comparable with control films where NGF was supplied via the medium. Future studies will determine the effect of typical neural prosthetic stimulation regimes on the release of neurotrophins and subsequent cell response.     

Kinetische und morphologische untersuchungen zur polymerisation von oxacyclischen monomeren im festen zustand, 2. Zur morphologie der bei der straheleninduzierten polymerisation von trioxan und tetroxan entstehenden poly(oxymethylene)

In order to clarify the reaction mechanism of the solid-state polymerisation of 1,3,5-trioxane and 1,3,5,7-tetroxane as induced by gamma-radiation, the nascent textures and morphology of poly(oxymethylene) crystals were studied with light and transmission electron microscopy, X-ray scattering and differential thermal analysis techniques. In both cases poly(oxymethylene) crystals with fibrous morphology and “twin-structure” are obtained. The polymerization of 1,3,5,7-tetroxane gives rise to the formation of crystals where the preferred orientation of the polymer chain is the monomer b-axis; however, the fibrils showed ripples perpendicular to the main chain direction at a 70–240Å long period that depends extremely on the polymerization temperature. In the polymerization of 1,3,5-trioxane, POM crystals with 60% of the polymer chains in the direction of the monomer c-axis are obtained. The remaining chains are found in the “twin sub-crystals”. Dark field electron micrographs showed the sub-crystals to be an integral part of the fibrils. The material in the twin direction decreases, when the polymerization temperature or the yields increases; it never amounts to less than 30%. These results and the misfit between the lattices of the monomers and the nascent polymer indicate that the reactions which take place occur according to a simultaneous polymerization and crystallization mechanism through the gasphase. The polymerization process occurs easier at higher temperatures near the monomers' melting point due to the increase of monomer vapor pressure and diffusion processes in the monomer crystals. Molecular models of the chain growth are developed on the basis of the morphological observations.     

Effect of suspension medium on the cationic polymerization of trioxane in solid state

Polyoxymethylene having high molecular weight was obtained by the solid state polymerization of trioxane crystals catalyzed by BF₃ · O(C₂H₅)₂, in a dispersing medium. To clarify the role of the medium, cyclohexane, hexane benzene and hexane-nitrobenzene were used. Addition of a small amount of good solvent increased the molecular weight of the resulting polymer, but the addition of a large amount of good solvent decreased molecular weight. When powdered crystals of trioxane were used in the solid state polymerization in n-hexane, both the rate of polymerization and molecular weight of polymer were larger than when large crystals were used. From these results, it is concluded that the first effect of a medium is the penetration into the crystal of monomer. This brings about an effective increase of both crystalline surface area and the easiness of diffusion of catalysts. The second effect of a medium is to protect the polymer from degradation by acid catalysts. If no liquid medium was used in solid state polymerization, high polymer could not be obtained at high conversion. The effect of a small amount of water was studied. The effect of water contained in the trioxane crystals was smaller than that of water in the liquid medium.     

Pop-corn polymerization of 2-hydroxyethyl methacrylate and the influence of a charged comonomer

The three-dimensional copolymerization of 2-hydroxyethyl methacrylate and ethylene dimethacrylate in the presence of water has been studied. An increasing concentration of ethylene dimethyacrylate results in the formation of a pop-corn polymer. The flakes of the pop-corn polymer initiate the polymerization of an added monomer which again results in a pop-corn polymer. The addition of a small percentage of charged monomer into the copolymerization batch (potassium methyacrylate) prevents the formation of a pop-corn polymer. An experimental investigation of the free-radical polymerization kinetics was carried out.     

Analysis of a dimethacrylate copolymer (bis-GMA and TEGDMA) network by DSC and 13C solution and solid-state NMR spectroscopy

In this work the effect of dilution with TEGDMA on the kinetics of Bis-GMA polymerization and on the extent of polymerization or degree of conversion was studied using (a) DSC and (b) NMR. The systems with lower viscosity and lower Tg exhibited higher extent of polymerization. For Bis-GMA/TEGDMA mixtures the calculated Tg values were found to be higher than the experimental values suggesting that a dilution effect is predominant rather than intermolecular hydrogen bonding. Solid state NMR has been shown to be a convenient method for measuring the total amount of conversion in a mixed monomer system. The disappearance of the NMR solution spectrum was used to reveal overall polymerization kinetics.     

Cell attachment functionality of bioactive conducting polymers for neural interfaces

Bioactive coatings for neural electrodes that are tailored for cell interactions have the potential to produce superior implants with improved charge transfer capabilities. In this study synthetically produced anionically modified laminin peptides DEDEDYFQRYLI and DCDPGYIGSR were used to dope poly(3,4-ethylenedioxythiophene) (PEDOT) electrodeposited on platinum (Pt) electrodes. Performance of peptide doped films was compared to conventional polymer PEDOT/paratoluene sulfonate (pTS) films using SEM, XPS, cyclic voltammetry, impedance spectroscopy, mechanical hardness and adherence. Bioactivity of incorporated peptides and their affect on cell growth was assessed using a PC12 neurite outgrowth assay. It was demonstrated that large peptide dopants produced softer PEDOT films with a minimal decrease in electrochemical stability, compared to the conventional dopant, pTS. Cell studies revealed that the YFQRYLI ligand retained neurite outgrowth bioactivity when DEDEDYFQRYLI was used as a dopant, but the effect was strongly dependant on initial cell attachment. Alternate peptide dopant, DCDPGYIGSR was found to impart superior cell attachment properties when compared to DEDEDYFQRYLI, but attachment on both peptide doped polymers could be enhanced by coating with whole native laminin.     

Early Stages of Propylene Bulk Phase Polymerization with Supported Metallocene Catalysts

For the initial steps of propylene bulk phase polymerization with a silica‐supported metallocene/MAO‐catalyst the processes of polymer growth, particle expansion and carrier fragmentation are investigated. Gravimetric analysis of the kinetics is given. Light optical examination of the particles shows a significant particle expansion during the first minutes of polymerization, which is not comparable to known observation from the slurry process. Electron microscopic investigation on cross sections of the polymer particles allows a detailed insight into the process of polymer growth, carrier fragmentation and particle expansion. The results are compared with the polymer growth and particle expansion model developed with slurry polymerizations under mild model conditions. In bulk phase polymerization particle expansion and carrier fragmentation are much faster due to more drastic reaction conditions. Nevertheless, it is still possible to observe a shell‐by‐shell degradation of the carrier material. Varying start up behaviors of single catalyst grains are observed. We are able to correlate the initial polymerization activity of individual grains with the cocatalyst loading and distribution on the catalyst carrier by means of microanalytical techniques. The results explain the varying start up behaviors of single grains. It becomes clear why different states of fragmentation are mixed up in the observed polymer samples.

Insight into catalyst grains after 0.5 min propylene bulk phase polymerization.