Retrograde labeling in peripheral nerve research: it is not all black and white.
ABSTRACT Retrograde labeling has become an important method of evaluation for peripheral nerve regeneration after injury. We review the features of the commonly used retrograde tracers Fast Blue, Fluoro-Gold, and Fluoro Ruby in addition to the various application methods (conduit reservoir, intramuscular injection, and crystal powder application) and the techniques used to count stained neurons. Upon application of the staining techniques and dyes in a rat and mouse nerve injury model, Fluoro-Gold was found to stain the greatest number of neurons with all application methods. However, due to variability of staining intensity, neuron size, and background staining, it is difficult to count the stained neurons accurately. Fast Blue stains consistently using intramuscular injection in the mouse but fails to provide adequate staining using the muscle injection method in the rat model and shows high failure rates using the conduit reservoir technique. However, crystal dye application with Fast Blue to the cut nerve end provides excellent results. We believe that it is imperative to use the various tracers and application methods prior to their experimental use to develop a consistent standardized approach to retrograde labeling.
- International Journal of Oral and Maxillofacial Surgery - INT J ORAL MAXILLOFAC SURG. 01/2011; 40(10):1135-1136.
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ABSTRACT: A drawback of electrical stimulation for muscle control is that large, fatigable motor units are preferentially recruited before smaller motor units by the lowest-intensity electrical cuff stimulation. This phenomenon limits therapeutic applications because it is precisely the opposite of the normal physiological (orderly) recruitment pattern; therefore, a mechanism to achieve orderly recruitment has been a long-sought goal in physiology, medicine and engineering. Here we demonstrate a technology for reliable orderly recruitment in vivo. We find that under optical control with microbial opsins, recruitment of motor units proceeds in the physiological recruitment sequence, as indicated by multiple independent measures of motor unit recruitment including conduction latency, contraction and relaxation times, stimulation threshold and fatigue. As a result, we observed enhanced performance and reduced fatigue in vivo. These findings point to an unanticipated new modality of neural control with broad implications for nervous system and neuromuscular physiology, disease research and therapeutic innovation.Nature medicine 10/2010; 16(10):1161-5. · 27.14 Impact Factor
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ABSTRACT: In the present study, the labeling efficacy of tracers Fluoro-ruby (FR), Fluoro-emerald (FE), True Blue (TB), Fluoro-Gold (FG), Diamidino Yellow (DY) and 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) to retrogradely label the cutaneous afferent neurons in the rat was examined. The proximal stump of the transected sural nerve was exposed for 1 hour either to one of the examined dyes (FR, FE, TB, FG, DY and DiI group) in single labeling experiments, or to mixtures of two dyes (TB-FG, FG-DiI, TB-DY and TB-DiI group) in double labeling experiments (n=5 for each group). After 10 days, dorsal root ganglia (DRGs) L3-S1 were harvested, cut to 20 microm thick longitudinal sections and all labeled neurons were counted. The average numbers of labeled DRG neurons in FR group (1063+/-158; mean+/-SD) and FE group (1067+/-203) were statistically significantly lower than those in TB group (2831+/-379), FG group (2802+/-134), DY group (2888+/-262) or DiI group (2900+/-278) (p<0.05). In double labeling experiments, the average number of double labeled neurons in TB-DY group (2208+/-207) was statistically significantly lower than those in TB-FG group (2775+/-316), FG-DiI group (2921+/-419), or TB-DiI group (2805+/-179) (p<0.05). Among examined tracers, TB, FG, DY and DiI, have the highest and similar labeling efficacy for retrograde labeling of cutaneous afferent neurons in the rat. The tracers TB, DiI and FG effectively label the same neuronal population in double labeling, therefore, their combinations are most suitable for double retrograde labeling studies of cutaneous afferent neurons in the rat.Journal of neuroscience methods 08/2010; 191(2):208-14. · 2.30 Impact Factor