Temporal interactive response is resistant to cloudy ocular media in the slow double-stimulation multifocal electroretinogram.

AIM: To examine the influence of cloudy media on the slow double-stimulation multifocal electroretinogram (mfERG). METHODS: Slow double-stimulation mfERG responses were measured from 26 subjects with normal ocular health under normal and light scattering conditions (induced using acrylic sheets) (Experiment 1) and another nine cataract patients before and after cataract surgery (Experiment 2). The amplitudes and implicit times of the first (M(1)) and second (M(2)) stimulation were compared under normal and light scattering conditions in Experiment 1 and they were compared under precataract and postcataract surgery in Experiment 2. RESULTS: Compared with control conditions (normal and postcataract surgery), the M(1) amplitude in the central region was significantly reduced in light scattering conditions (acrylic sheets and precataract surgery); the M(2) amplitude and both M(1) and M(2) implicit times of all regions examined were moderately affected in precataract surgery. The M(1):M(2) amplitude ratio and implicit time ratio were virtually unaffected in cloudy media for either central or mid-peripheral regions. CONCLUSION: Cloudy media affects the mfERG amplitude and implicit time in the slow double-stimulation, but does not affect the response ratio (ie, M(1):M(2) amplitude ratio and implicit time ratio) between the two stimulations. This suggests that the ratio analysis can be applied in patients with mild to moderately cloudy ocular media to evaluate the functional integrity of the retina.

Luminance-modulated adaptation in the global flash mfERG: a preliminary study of early retinal functional changes in high-risk glaucoma patients.

PURPOSE: To investigate the association of the luminance-modulation global flash multifocal electroretinogram (mfERG) and other clinical assessments of vision in subsets of subjects at high risk of developing glaucomatous damage. METHODS: Eighteen subjects (28 eyes) with asymmetric glaucoma and ocular hypertension were measured in this longitudinal study of visual field, OCT, and multifocal electroretinogram (mfERG). Five ophthalmic examinations were scheduled, once every 12 months over a 4-year period. The mfERG was assessed using a luminance-modulated global flash stimulation paradigm. The adaptive index which we have reported previously was calculated. RESULTS: There was a significant thinning of the peripapillary retinal nerve fiber layer over the course of the study for eyes with ocular hypertension, or for fellow eyes with asymmetric glaucoma which initially had an abnormal adaptive index; such eyes showed a thinning rate of -3.59 and -3.69 µm/year, respectively. However, no significant thinning was found for eyes which initially had a normal adaptive index. Two subjects were shown to have glaucomatous damage, confirmed by abnormal thinning of the retinal nerve fiber layer and visual field loss respectively at the last visit. However, these patients had shown an abnormal adaptive index in the mfERG measurement at the first visit. CONCLUSIONS: The adaptive index calculated from the measurement of luminance-modulated global flash mfERG is useful for predicting progression of signs related to glaucoma, especially in high-risk groups. The abnormal adaptive index reflects the change in fast-adaptive mechanisms in the retina and indicates the risk of developing glaucoma.

Effect of inner retinal dysfunction on slow double-stimulation multifocal electroretinogram.

PURPOSE: This study investigated the retinal adaptive mechanism in inner retinal dysfunction using the slow double-stimulation multifocal electroretinogram (mfERG) paradigm. METHODS: Slow double-stimulation mfERG responses were recorded from 15 eyes of 15 4-month-old Mongolian gerbils in control conditions and after suppression of inner retinal responses with injections of tetrodotoxin (TTX) and N-methyl-d-aspartic acid (NMDA). The stimulation consisted of five video frames: the two initial frames with multifocal flashes were triggered by two independent m-sequences, followed by three dark video frames. The results were compared with findings in humans: 7 subjects with glaucoma and 31 age-matched normal subjects were measured using the same mfERG protocol. RESULTS: The stimulation generates two responses (M(1) and M(2)) from the two independent multifocal frames. The M(1):M(2) ratio showed a significant reduction after administration of TTX+NMDA in the animal study. This matched with the human glaucoma findings. Glaucoma subjects generally have a reduced M(1):M(2) ratio; this ratio showed a sensitivity of 86%, with a specificity of 84% for differentiating normal eyes from glaucomatous eyes. CONCLUSION: This stimulation paradigm provides a method of measuring temporal visual characteristics. The M(1):M(2) ratio acts as an indirect functional indicator of retinal adaptation, which may be abnormal in the diseased retina. Further development of this method may help to describe the functional variation in the diseased retina and to predict the occurrence of a range of retinopathies.

Forward and backward adaptive effects in global flash multifocal electroretinogram stimulation.

PURPOSE: The present study investigated retinal adaptive responses in concert with the modulation of forward and backward adaptation induced by periodic global flashes using the global flash multifocal electroretinogram (mfERG). METHODS: Six normal subjects were recruited for global flash mfERG measurements, which consisted of 103 scaled hexagonal elements followed by a global flash frame. In experiments I and II, with constant luminance maintained in both local and global flash frames, the number of dark frames was independently varied and these frames were either inserted prior to or following the global flash frame to investigate the forward or backward adaptive effect of the global flash on the mfERG. In experiment III, the number of dark frames was fixed but the luminance of the global flash frame was varied with constant luminance of the focal flash. This was used to demonstrate that the adaptive effect related not to time but to variation of luminance. RESULTS: All the central, para-central and peripheral direct component amplitudes were found to be significantly influenced by variation of the number of dark frames (p < 0.01). Reducing the forward adaptive effect of the global flash enhanced the direct component response and it became steady after five dark frames were inserted following the global flash. Reducing the backward adaptive effect of the global flash also enhanced the direct component response but it started reducing after four dark frames were inserted prior to the global flash frame. These changes were different with luminance modulation of the global flash intensity with fixed dark frames, while the direct component amplitude grew approximately linearly with decreasing mean luminance of the global flash stimulation. CONCLUSION: The retina plays a major role in visual adaptation. Both forward and backward adaptive effects of the global flash on the direct component have been illustrated in this study. The results show that the forward and backward adaptive phenomena in the global flash mfERG are different and demonstrate that backward adaptation is found at the retinal level.

Detection of early functional changes in diabetic retina using slow double-stimulation mfERG paradigm.

Aim Diabetes mellitus (DM) is a systemic disease with insufficient secretion of insulin or poor response to insulin. This typically causes poor control of blood glucose level leading to a range of complications. Early detection of the retinal function alteration in DM is needed. Methods A newly modified paradigm-slow double-stimulation multifocal electroretinogram (mfERG)-was introduced to measure early changes of retinal function in DM and to investigate changes in the adaptation mechanisms in the diabetic retina. The mfERG was measured by using a slow double-stimulation mfERG paradigm (M(1)M(2)OOO). Results The m1 amplitude of M(1) stimulation from diabetic subjects was significantly reduced in ring 1 in contrast to that of a control group. The m2 amplitude of M(2) stimulation from diabetic subjects was also significantly reduced in ring 1 and 2 as compared with those of the control group. The m1/m2 ratio which minimises intersubject variation shows a reasonable differentiation between the control and diabetic groups. There was a significant increase in the amplitude ratio from diabetic subjects in ring 2 and 3 as compared with those of the control group. Conclusions The present findings suggest that the new mfERG paradigm is a fast and sensitive test for the detection of early functional changes in the diabetic retina.

Impairment of retinal adaptive circuitry in the myopic eye.

Previous studies have proposed that the inner retina is affected in myopes. This study aimed to investigate the changes in adaptive circuitry of the inner retina in myopia, using the global flash multifocal electroretinogram (global flash mfERG) with different levels of contrast (luminance modulation). Fifty-four myopes had global flash mfERG recorded with different contrasts. The direct component (DC) and the induced component (IC) of the mfERG response were pooled into six regions for analysis. The response amplitudes and implicit times at different contrasts were also analysed. Results showed that myopes had significant reduction in the paracentral DC amplitude for the 29% and 49% contrasts and in the paracentral IC amplitude at all contrasts measured. The peripheral IC amplitude for the 49% contrast was also reduced. No significant change was found in implicit time for either DC or IC response. Refractive error explained about 14% of the variance in DC and 16% of the variance in IC amplitude respectively; axial length could not account for additional variance in either paracentral DC or IC amplitudes in the hierarchical regression models used. We concluded that the paracentral retinal region in myopes showed signs of impaired retinal adaptation, suggesting a functional loss at the inner retinal layer. In addition, functions attributed to the outer retinal layer showed only small changes due to myopia.

Porcine global flash multifocal electroretinogram: possible mechanisms for the glaucomatous changes in contrast response function.

PURPOSE: The aim of this study was to obtain a better understanding of the cellular contributions to the porcine global flash mfERG by using a pharmacologic dissection method, together with the method using variation of stimulus contrast which has been used to demonstrate mfERG changes in human glaucoma. METHODS: Global flash mfERGs with different stimulus-contrast settings (99%, 65%, 49% or 29%) were recorded from 14 eyes of ten 6-week-old Yorkshire pigs in control conditions and after suppression of inner retinal responses with inhalation of isoflurance (ISO), and injections of tetrodotoxin (TTX) and N-methyl-d-aspartic acid (NMDA). ON- and OFF-pathway responses were isolated by injection of 2-amino-4-phosphonobutyric acid (APB) and cis-2,3-piperidinedicarboylic acid (PDA). RESULTS: The porcine global flash mfERG consisted of an early direct component (DC) and a late induced component (IC). ISO and TTX removed inner retinal contributions to the IC; NMDA application further abolished the oscillatory wavelets in the DC and removed the residual IC waveform. The inner retina contributed regular oscillation-like wavelets (W1, W2 and W3) to the DC and shaped the IC. After removing the inner retinal contributions, the porcine global flash mfERG waveform becomes comparable to that obtained with conventional mfERG stimulation. The remaining waveform (smoothed DC) was mainly contributed by the ON- and OFF-bipolar cells as revealed after APB or PDA injection. Photoreceptors contributed a small signal to the leading edge of N1. The characteristic of contrast response function of DC was demonstrated to be contributed by the inner retinal oscillation-like wavelets. CONCLUSION: We believe that the DC of the porcine global flash mfERG is mainly composed of contributions from photoreceptors, and ON- and OFF-bipolar cells, where inner retinal activity partially shaped the DC with superimposed regular wavelets. However, the IC is dominated by inner retinal activity. The contrast response functions of DC consisted of both outer retinal response and oscillation-like wavelets of the inner retinal response. Both contain different characteristics during contrast modulation of the stimulus, where the changes of W2 of the inner retinal response seem independent of contrast modulation. The DC contrast response feature depends mainly on the relative contribution of inner retinal activities; the loss of inner retinal cells may alter the DC contrast response function, making it tend toward linearity.

The characteristics of multifocal electroretinogram in isolated perfused porcine eye: cellular contributions to the in vitro porcine mfERG.

We investigated characteristics of multifocal electroretinograms (mfERG) from in vitro perfused porcine eyes. TTX, NMDA, APB, and PDA were used to identify contributions to the mfERG from inner retinal neurons, ON-pathway, OFF-pathway, and photoreceptors. The cellular contributions of the first-order kernel (K1) in an isolated perfused porcine mfERG came from both inner and outer retina, and were similar to those of in vivo porcine mfERG. In addition, application of isoflurane and propofol caused interference with the mfERG response which resembled inner retinal activities sensitive to TTX + NMDA application. Improved understanding of the cellular origins of the perfused porcine mfERG, in the absence of anesthetic agents, is useful for identifying changes shown in the waveform under anesthesia.

Multifocal electroretinogram in rhodopsin P347L transgenic pigs.

PURPOSE: Neural ectopic rewiring in retinal degeneration such as retinitis pigmentosa (RP) may form functional synapses between cones and rod bipolar cells that cause atypical signal processing. In this study, the multifocal electroretinograms (mfERGs) of a large animal model of RP, the rhodopsin P347L transgenic (Tg) pig, were measured to examine the sources and nature of altered signal processing. METHODS: mfERG responses from a 6-week-old Tg pig were recorded before and after sequential application of tetrodotoxin (TTX), N-methyl-D-aspartate (NMDA), 2-amino-4-phosphonobutyric acid (APB), and cis-2,3-piperidinedicarboylic acid (PDA), to identify contributions to the retinal signal from inner retinal neurons, the ON-pathway, the OFF-pathway, and photoreceptors. The mfERG response contributions from different retinal components of in the Tg eyes were estimated and compared with control data from eyes of age-matched wild-type (WT) pigs. RESULTS: There was a prominent difference in the estimates of the inner retinal response and ON-bipolar cell pathway contribution between the Tg and WT mfERG responses. In particular, the early components of the inner retinal contribution were obviously altered in the Tg mfERG. The inner retinal components at approximately 24 and 40 ms appeared to be inverted. Differences in the estimates of OFF-bipolar cell pathway contributions were minimal. There was no change in cone cell responses in the Tg mfERG. CONCLUSIONS: In Tg retinas, ectopic synapses formed between cones and rod bipolar cells probably altered signal processing of the ON-bipolar cell pathway. In response to the altered visual signal input from the outer retina, signal processing in inner retinal neurons was also modified.

Pharmacologically defined components of the normal porcine multifocal ERG.

Multifocal electroretinograms (mfERG) from isoflurane anesthetized pigs were recorded and sequential application of TTX, NMDA, APB and PDA were used to identify contributions to the mfERG from inner retinal neurons, ON-pathway, OFF-pathway and photoreceptors. The cellular origins of the first-order kernel (K1) and the first slice of the second-order kernel (K2.1) porcine mfERG are contributed from both inner and outer retina. For the K1 waveform, the n1 involved responses of cone photoreceptors and OFF-bipolar cells. The leading edge of p1 is dominated by ON-bipolar cell depolarization. The rear edge of p1, n2 and p2 are dominated by ON-bipolar activities and shaped by the activities of OFF-bipolar cells and retinal cells with NMDAr and voltage-gated sodium channels other than ganglion cells. The p3 is mainly inner retinal activities. For the K2.1 waveform, the p1 and n1 are the summation of activities of ON-, OFF-bipolar cells and retinal cells rich in NMDAr and voltage-gated sodium channels other than ganglion cells. The p2 seems to be related to the ganglion cells. Better understanding of the cellular origins of the normal porcine mfERG will be useful for comparing and defining the functional changes that may occur in diseased retinas.

Construction of multiporphyrin arrays using ruthenium and rhodium coordination to phosphines.

The synthesis of linear multiporphyrin arrays with mono- and bisphosphine-substituted porphyrins as ligand donors and ruthenium(II) or rhodium(III) porphyrins as ligand acceptors is described. With appropriate amounts of the building blocks mixed, linear dimeric and trimeric arrays have been synthesized and analyzed by (1)H NMR and (31)P NMR spectroscopy. The Ru/Rh acceptor porphyrins can be located either at the periphery or in the center of the array. Likewise, the monophosphine porphyrins can be positioned at the periphery, thus allowing a high degree of freedom in the overall composition of the arrays. This way, both donor and acceptor porphyrins can act as chain extenders or terminators. One of the trimeric complexes with two nickel and one ruthenium porphyrin has also been analyzed by X-ray crystallography. Attempts have also been made to synthesize higher order arrays by mixing appropriate amounts of the porphyrins; however, from the NMR data it cannot be concluded if monodisperse five, seven, or nine porphyrin arrays are present or if the solutions are composed of a statistical mixture of smaller and larger arrays.

Phosphine and phosphonite complexes of a ruthenium(II) porphyrin. 1. Synthesis, structure, and solution state studies.

We have investigated the effect of complexation of different phosphorus ligands on the stability, solid state structure, and spectroscopic properties (NMR, IR, UV-vis) of a 5,15-diphenyl-substituted ruthenium porphyrin, (MeOH)Ru(II)(CO)(DPP) 2 [DPP = 5,15-bis(3',5'-di-tert-butyl)phenyl-2,8,12,18-tetraethyl-3,7,13,17-tetramethylporphyrin]. The ligands used are PPh(3), diphenyl(phenylacetenyl)phosphine (DPAP), bis(diphenylphosphino)acetylene (DPPA), tris(phenylacetenyl)phosphine [(PA)(3)P], and diethyl (phenylacetenyl)phosphonite [PAP(OEt)(2)]. The mono-phosphine complexes (PR(3))Ru(II)(CO)(DPP) are readily formed in solution in quantitative yields. The complexes display association constants ranging from 1.2 x 10(4) M(-1) for PPh(3) to 4.8 x 10(6) M(-1) for PAP(OEt)(2). The weak association of PPh(3) does not correlate with its pK(a), delta((31)P), or cone angle value and is attributed to steric effects. Due to their kinetic lability, which is shown by 2D NMR spectroscopy, and the weakening of the carbonyl ligand via a trans effect, the mono-phosphine complexes could not be isolated. IR spectroscopy gives the relative order of pi-acceptor strength as PPh(3) < DPAP, DPPA < (PA)(3)P < PAP(OEt)(2), whereas the relative order of the sigma-donor strength is PPh(3) < (PA)(3)P < DPAP, DPPA < PAP(OEt)(2), based on the calculated pK(a) values and on the (31)P((1)H) NMR chemical shifts of the ligands. The chemical shift differences in the (31)P9(1)H)) NMR spectra upon ligand binding display a linear correlation with the calculated pK(a) values of the protonated ligands HPR(3)(+); we propose that the pK(a), and probably other electronic properties, of a specific phosphorus ligand can be estimated on the basis of the chemical shift difference Deltadelta((31)P) upon complexation to a metalloporphyrin. The bis-phosphine complexes can be isolated in pure form by crystallization from CHCl(3)-MeOH solutions using excess ligand. Association of the second ligand is in the same order of magnitude as the first binding for the phosphines, but the second phosphonite binding is decreased by a factor of about 100. The solid state structures show only marginal differences in the geometrical parameters. The calculated and the crystallographic cone angles of the ligands generally do not match, apart from the values obtained for PAP(OEt)(2).

Amplification of a cyclic mixed-metalloporphyrin tetramer from a dynamic combinatorial library through orthogonal metal coordination.

A cyclic porphyrin tetramer, consisting of two bis-phosphine substituted zinc(II) porphyrin units and two Rh(III)TPP units, is selected and amplified virtually quantitatively from a dynamic combinatorial library using 4,4'-bipy as a scaffold and using orthogonal binding modes.

Identification and Isolation of a Receptor for N-Methyl Alkylammonium Salts: Molecular Amplification in a Pseudo-peptide Dynamic Combinatorial Library.

A cyclic pseudo-peptide receptor for acetylcholine has been amplified and isolated from a dynamic combinatorial library by virtue of templated stabilization under thermodynamic control (see scheme, TFA=trifluoroacetic acid). This is a demonstration of significant molecular amplification in dynamic systems to evolve a novel receptor.