Benedetto Falsini

UV- and midwave-sensitive cone-driven retinal responses of the mouse: A possible phenotype for coexpression of cone photopigments

Molecular biological, histological and flicker electroretinographic results have established that mice have two cone photopigments, one peaking near 350 nm (UV-cone pigment) and a second near 510 nm [midwave (M)-cone pigment]. The goal of this investigation was to measure the action spectra and absolute sensitivities of the UV-cone- and M-cone-driven b-wave responses of C57BL/6 mice. To achieve this goal, we suppressed rod-driven signals with steady or flashed backgrounds and obtained intensity–response relations for cone-driven b-waves elicited by narrowband flashes between 340 and 600 nm. The derived cone action spectra can be described as retinal1pigments with peaks at 355 and 508 nm. The UV peak had an absolute sensitivity of ∼8 nV/(photon μm2) at the cornea, approximately fourfold higher than the M peak. In an attempt to isolate UV-cone-driven responses, it was discovered that an orange conditioning flash (λ > 530 nm) completely suppressed ERG signals driven by both M pigment- and UV pigment-containing cones. Analysis showed that the orange flash could not have produced a detectable response in the UV-cone pathway were their no linkage between M pigment- and UV pigment-generated signals. Because cones containing predominantly the UV and M pigments have been shown to be located largely in separate parts of the mouse retina (Szel et al., 1992), the most probable linkage is coexpression of M pigment in cones primarily expressing UV pigment. New histological evidence supports this interpretation (Gloesman and Ahnelt, 1998). Our data are consistent with an upper bound of ∼3% coexpression of M pigment in the cones that express mostly the UV pigment.

Visually evoked hemodynamical response and assessment of neurovascular coupling in the optic nerve and retina

The retina and optic nerve are both optically accessible parts of the central nervous system. They represent, therefore, highly valuable tissues for studies of the intrinsic physiological mechanism postulated more than 100 years ago by Roy and Sherrington, by which neural activity is coupled to blood flow and metabolism. This article describes a series of animal and human studies that explored the changes in hemodynamics and oxygenation in the retina and optic nerve in response to increased neural activity, as well as the mechanisms underlying these changes. It starts with a brief review of techniques used to assess changes in neural activity, hemodynamics, metabolism and tissue concentration of various potential mediators and modulators of the coupling. We then review: (a) the characteristics of the flicker-induced hemodynamical response in different regions of the eye, starting with the optic nerve, the region predominantly studied; (b) the effect of varying the stimulus parameters, such as modulation depth, frequency, luminance, color ratio, area of stimulation, site of measurement and others, on this response; (c) data on activity-induced intrinsic reflectance and functional magnetic resonance imaging signals from the optic nerve and retina. The data undeniably demonstrate that visual stimulation is a powerful modulator of retinal and optic nerve blood flow. Exploring the relationship between vasoactivity and metabolic changes on one side and corresponding neural activity changes on the other confirms the existence of a neurovascular/neurometabolic coupling in the neural tissue of the eye fundus and reveals that the mechanism underlying this coupling is complex and multi-factorial. The importance of fully exploiting the potential of the activity-induced vascular changes in the assessment of the pathophysiology of ocular diseases motivated studies aimed at identifying potential mediators and modulators of the functional hyperemia, as well as conditions susceptible to alter this physiological response. Altered hemodynamical responses to flicker were indeed observed during a number of physiological and pharmacological interventions and in a number of clinical conditions, such as essential systemic hypertension, diabetes, ocular hypertension and early open-angle glaucoma. The article concludes with a discussion of key questions that remain to be elucidated to increase our understanding of the physiology of ocular functional hyperemia and establish the importance of assessing the neurovascular coupling in the diagnosis and management of optic nerve and retinal diseases.

Remodeling of second-order neurons in the retina of rd/rd mutant mice

This is a brief review of data obtained by analyzing the morphology and the physiology of the retinas in rd/rd and normal, wt mice, aged 10-90 days. Second-order neurons of the rd/rd show abnormalities that start with the anomalous development of rod bipolar cells around P10 and culminate with the atrophy of dendrites in cone bipolar cells, mostly evident at P90. Horizontal cells remodel considerably. Cone-mediated ERGs, (recorded between 13 and 16 days of age) have reduced a-wave and b-wave amplitudes and longer b-wave latency and duration. B-wave abnormalities indicate specific postreceptoral dysfunction. Morphological and ERG changes in rd/rd retinas are consistent with substantial inner retinal remodeling associated to photoreceptor degeneration.

Pattern electroretinogram as a function of spatial frequency in ocular hypertension and early glaucoma

Pattern electroretinograms (PERGs) in response to 8.3 Hz alternating gratings (16.6 rev/sec) of different spatial frequencies were recorded in normal subjects as well as in patients affected by early glaucoma and ocular hypertension. In normal subjects the PERG response is spatially tuned, with a maximum at about 1.5 cycles/deg and attenuation at higher and lower spatial frequencies. In all cases of early glaucoma and in the great majority of cases of ocular hypertension the PERG was reduced, as compared with that of normal subjects, mainly in the medium range of spatial frequencies (at which the PERG has its maximal amplitude).

The Origin of the Major Rod- and Cone-Driven Components of the Rodent Electroretinogram and the Effect of Age and Light-Rearing History on the Magnitude of These Components

The electroretinogram (ERG) is a complex field potential evoked by light. The ERG response to brief flashes has long provided an important assessment tool in basic and clinical research (see, e. g., Berson, 1992; Jacobson et al., 1994). In the past decade, several rapidly growing and overlapping areas of retinal research have led to extensive basic and clinical applications of the ERG.

Influence of short-term antioxidant supplementation on macular function in age-related maculopathy. A pilot study including electrophysiologic assessment

PURPOSE To evaluate the influence of short-term antioxidant supplementation on retinal function in age-related maculopathy (ARM) patients by recording focal electroretinograms (FERGs). DESIGN Nonrandomized, comparative clinical trial. PARTICIPANTS Thirty patients with early ARM and visual acuity >/=20/30, divided into two groups, similar for age and disease severity: antioxidant group (ARM-A, n = 17) and no treatment group (ARM-NT, n = 13). Eight age-matched normal subjects divided into antioxidant (N-A, n = 4) or no treatment (N-NT, n = 4) groups. METHODS ARM-A patients and N-A patients had oral supplementation of lutein, 15 mg; vitamin E, 20 mg; and nicotinamide, 18 mg, daily for 180 days, whereas ARM-NT patients and N-NT patients had no dietary supplementation during the same period. Eight of the 17 ARM-A patients took supplementation for an additional 180-day period. In all patients and normal subjects, FERG assessment was performed at the study entry (baseline) and after 180 days. Further testing was performed at 360 days for the eight ARM-A patients taking supplements and for one ARM-A patient who had discontinued supplementation after 180 days. FERGs were recorded in response to a 41-Hz sinusoidally modulated uniform field (93.5% modulation depth) presented to the macular region (18 degrees ) on a light-adapting background. In a subgroup of patients (11 ARM-A and 5 ARM-NT), whose responses had suitable signal-to-noise ratios, FERGs were also recorded at different stimulus modulation depths between 8.25% and 93.5%. MAIN OUTCOME AND MEASURES Amplitude (in micro V) and phase (in degrees) of the FERG fundamental harmonic component. FERG modulation thresholds, estimated from the value of log modulation depth yielding a criterion response. RESULTS At 180 days, FERGs of ARM-A patients and N-A patients were increased in amplitude (mean change, 0.11 and 0.15 log micro V, respectively, P </= 0.01) compared with baseline values, whereas no significant changes in FERG amplitudes of ARM-NT patients and N-NT patients were found (mean change, -0.004 and -0.023 log micro V, respectively). In all groups no changes in the FERG phase were found. FERG modulation thresholds decreased with respect to baseline values (mean change, -0.36 log units, P < 0.01) in ARM-A patients, whereas no significant change (mean change, 0.07 log units) in ARM-NT patients was seen. At 360 days, FERGs of ARM-A patients taking supplementation were still increased in amplitude with respect to baseline (P < 0.05) but did not differ from those recorded at 180 days. In the patient who had discontinued supplementation, FERG amplitude decreased from the 180 days value, approaching that recorded at baseline. CONCLUSIONS Although this study provides no evidence for the long-term benefit of antioxidants in ARM, the results suggest that increasing the level of retinal antioxidants might influence macular function early in the disease process, as well as in normal aging.

Nonselective Loss of Contrast Sensitivity in Visual System Testing in Early Type I Diabetes

Objective Psychophysical methods in patients with diabetes mellitus reveal deficits of central or foveal vision. Our aim was to evaluate the contrast-sensitivity thresholds in 24 insulin-dependent (type I) diabetic patients with a short disease duration and without retinopathy, taking into account metabolic control. Research Design and Methods The control group consisted of age-matched nondiabetic subjects. None had visual or systemic symptoms. Contrast sensitivity measured at eight different spatial frequencies to sinusoidal bar patterns of 0.6–12.2 cycles/deg can detect functional defects in the spatially sensitive retinal ganglion cells or in higher visual pathways. We performed two different temporal types of contrast-sensitivity testing, dynamic (8 Hz) and static (0 Hz). Results Significant losses with dynamic contrast-sensitivity test at all but the highest spatial frequencies (i.e., 12.2 cycles/deg) were shown, whereas there was significant attenuation of contrast sensitivity at five spatial frequencies (1.0, 1.4, 2.2, 7.1, and 9.6 cycles/deg) in the static mode. Grating losses (<2SD of control means) of contrast sensitivity were found in 33.3% (dynamic) and in 72.9% (static) of eyes of diabetic patients. HbA1c values were positively correlated at variable spatial frequencies (1.0, 1.4, and 2.2 cycles/deg for dynamic test and 0.6, 1.0, 1.4, 2.2, 4.8, and 7.1 cycles/deg for static test). Conclusions Our results suggest an early, generally nonselective neuronal damage of visual pathways that occurs before the onset of clinically detectable retinopathy. The visual deficit may be related directly to the effects of diabetes; repetitive minor hypoglycemic insults may contribute more than a marked hyperglycemic condition to the mechanisms underlying physiological changes along the optic nerve.

Intravitreal bevacizumab (Avastin®) in proliferative diabetic retinopathy

Purpose:  To evaluate the efficacy and safety of intravitreal bevacizumab in proliferative diabetic retinopathy (PDR) patients.

Subfoveal choroidal blood flow and central retinal function in retinitis pigmentosa

PURPOSE To determine whether subfoveal choroidal blood flow is altered in retinitis pigmentosa (RP) and whether this alteration is associated with central cone-mediated dysfunction. METHODS In 31 RP patients (age range, 15-72 years) with preserved visual acuity (range: 20/30-20/20), subfoveal choroidal blood flow was measured by real-time, confocal laser Doppler flowmetry, and focal macular (18°) electroretinograms (FERGs) were elicited by 41 Hz flickering stimuli. Twenty normal subjects served as controls. The following average blood flow parameters were determined based on three 60-second recordings: volume (ChBVol), velocity (ChBVel), and flow (ChBF), the last being proportional to blood flow if the hematocrit remains constant. The amplitude and phase of the FERG first harmonic component were measured. RESULTS On average, ChBF and ChBVel were reduced by 26% (P ≤ 0.02) in RP patients compared to controls, whereas ChBVol was similar in the two groups. FERG amplitudes were reduced by 60% (P < 0.01) in patients compared with controls. FERG phases of patients tended to be delayed (P < 0.08) compared with their values in the controls. In patients, FERG phase delays were correlated (r = 0.50, P < 0.01) with ChBF and ChBVel values. FERG amplitudes were correlated (r = 0.49, P < 0.01) with ChBVol values. CONCLUSIONS These data indicate significant alterations of subfoveal choroidal hemodynamic in RP and suggest a link between the alteration of ChBF and the RP-associated central cone-mediated dysfunction as assessed by the FERG.

Detection of inner retina dysfunction by steady-state focal electroretinogram pattern and flicker in early IDDM

The effects of diabetes on the neural retina before the onset of clinically detectable retinopathy can be investigated with electrophysiological methods. Our aim was to detect early retinal dysfunctions in 60 patients with insulin-dependent diabetes mellitus (IDDM) and with a short duration of disease. We used the steady-state focal (9° field size) electroretinogram (ERG) of the macula in response to luminance modulation of a uniform field (flicker ERG) or to counterphase-modulated sinusoidal gratings (pattern ERG). The harmonic analysis of flicker ERG and pattern ERG yielded three main components: a first and a second harmonic to flicker (1F and 2F, respectively) and a second harmonic to pattern (2P). The 1F is believed to be correlated to photoreceptor activity, whereas 2F and 2P represent different subsets of generators in the inner retina. Results of focal ERG in IDDM patients with no or early retinopathy were compared with age-matched control subjects. Mean 2F and 2P amplitudes were significantly reduced in IDDM patients compared with the control group (P = 0.0001 by analysis of variance). 2P but not 2F amplitude was significantly more reduced in patients with retinopathy than in those without retinopathy (P < 0.05). 2F but not 2P phase abnormalities were observed in some patients. 2F and 2P alterations were slightly correlated with metabolic control (r = 0.22, P = 0.02) and disease duration (r = 0.28, P = 0.003). 1F was not significantly altered in IDDM patients. Our results suggest that early diabetes causes selective neurosensory deficits of inner retina layers, whereas the photoreceptors appear unaffected.