Multiphoton fluorescence microscopy offers the advantages of deep optical sectioning of living tissue with minimal phototoxicity and high optical resolution. More importantly, dynamic processes and multiple functions of an intact organ can be visualized in real time using noninvasive methods, and quantified. These studies aimed to extend existing methods of multiphoton fluorescence imaging to directly observe and quantify basic physiological parameters of the kidney including glomerular filtration rate (GFR) and permeability, blood flow, urinary concentration/dilution, renin content and release, as well as more integrated and complex functions like the tubuloglomerular feedback (TGF)-mediated oscillations in glomerular filtration and tubular flow. Streptozotocin-induced diabetes significantly increased single-nephron GFR (SNGFR) from 32.4 +/- 0.4 to 59.5 +/- 2.5 nl/min and glomerular permeability to a 70-kDa fluorophore approximately eightfold. The loop diuretic furosemide 2-fold diluted and increased approximately 10-fold the volume of distal tubular fluid, while also causing the release of 20% of juxtaglomerular renin content. Significantly higher speeds of individual red blood cells were measured in intraglomerular capillaries (16.7 +/- 0.4 mm/s) compared with peritubular vessels (4.7 +/- 0.2 mm/s). Regular periods of glomerular contraction-relaxation were observed, resulting in oscillations of filtration and tubular flow rate. Oscillations in proximal and distal tubular flow showed similar cycle times ( approximately 45 s) to glomerular filtration, with a delay of approximately 5-10 and 25-30 s, respectively. These innovative technologies provide the most complex, immediate, and dynamic portrayal of renal function, clearly depicting the components and mechanisms involved in normal physiology and pathophysiology.
McCulloch, Fiona, Régine Chambrey, Dominique Eladari, and János Peti-Peterdi. Localization of connexin 30 in the luminal membrane of cells in the distal nephron.
The aim of this study was to assess the effect of potential target tissue on regenerating neurones of the snail Lymnaea stagnalis using the three-dimensional collagen gel culture system. Mammalian type I collagen supported the regenerative outgrowth of snail neurones, and the neurofilament antibody SMI31 specifically labelled regenerating processes both within the gel and those growing over the surface of the ganglia. Using these techniques we tested the effect of co-culturing ganglia with either additional nervous tissue, previously shown to produce trophic substances, or buccal muscle on both the amount and direction of outgrowth. We conclude that, under the conditions used, neither target tissue provided trophic or tropic support in collagen gel cultures.
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