Electroretinographic assessment of rod- and cone-mediated bipolar cell pathways using flicker stimuli in mice.

Mouse full-field electroretinograms (ERGs) are dominated by responses of photoreceptors and depolarizing (ON-) bipolar cells, but not much of hyperpolarizing (OFF-) bipolar cells under conventional recording conditions. Here we investigate a novel ERG protocol in mice for functional assessment of the major ON- and OFF-bipolar cell pathways using flicker stimuli for a high luminance with varying frequency up to 30 Hz. Wild-type (WT) and functionally specific transgenic mice (Cnga3(-/-), no cone photoreceptor function; rho(-/-), no rod photoreceptor function; mGluR6(-/-), no ON-bipolar cell function) were examined. The Cnga3(-/-) flicker ERG was similar to the WT flicker ERG at very low stimulus frequencies, whereas ERGs were comparable between WT and rho(-/-) mice at 5 Hz and above. Between 5 and 15 Hz, ERGs in mGluR6(-/-) mice differed in configuration and amplitude from those in WT and rho(-/-) mice; in contrast, response amplitudes above 15 Hz were comparable among WT, rho(-/-) and mGluR6(-/-) mice. In summary, we found three frequency ranges with these conditions that are dominated by activity in the rod pathways (below 5 Hz), cone ON-pathway (between 5 and 15 Hz), and cone OFF-pathway (above 15 Hz) that enables a quick overview of the functionality of the major bipolar cell pathways.

Size-selective and in vitro assessment of inner blood retina barrier permeability.

Assessment of tight junction integrity in vitro is fundamental when studying molecular processes that may be implicated in barrier dysfunction. At the blood brain and inner blood retina barrier (BBB and iBRB, respectively) adjacent endothelial cells lining the microvasculature have been shown to have very low rates of fluid phase transcytosis and high electrical resistances, due in part to the expression of tight junction proteins at the apical periphery of these cells. While these high electrical resistances are difficult to achieve in vitro, owing to complex interactions of endothelial cells in vivo with astrocytes and pericytes, it is possible to make an assessment of paracellular permeability when cells are analysed on a number of different fronts. In this regard, we will outline here a method for determining trans-endothelial electrical resistance, tracer molecule diffusion, and tight junction protein localization in primary cultures of bovine retinal microvascular endothelial cells. This system allows for the screening of a wide range of pro- and anti-angiogenic molecules in an in vitro model of the iBRB and can accurately assess the role individual tight junction proteins play in maintaining tight junction integrity in response to various cell stimuli.

From RNA interference technology to effective therapy: how far have we come and how far to go?

Over a decade has passed since the first description of RNAi in animals--the fundamental endogenous process by which small dsRNAs mediate sequence-specific gene silencing. This discovery has radically transformed our understanding of gene regulation and function and spawned a whole new biotechnology industry focused on developing RNAi-based therapeutic approaches to a variety of human diseases that have otherwise proved challenging to conventional therapies. While RNAi technologies hold great promise as a powerful medical tool, successful delivery of RNAi agents and effective measurement of their uptake are major challenges in translating RNAi therapies to the clinic. Exciting developments in the field have also been tempered by safety concerns surrounding the immunogenic potential of this gene silencing technology and the potential side effects associated with exploiting a crucial biological pathway for therapeutic benefit. This article examines the progress of RNAi therapeutics including advances in delivery and safety, and recent findings from several Phase I-III clinical trials. The emergence of a novel application of RNAi in enhancing the delivery of low-molecular weight drugs to neuronal tissues will also be presented to provide an outlook on the future of RNAi technologies.