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After reinnervation following transection of the sciatic nerve, normal patterns of regional type I fiber distribution are known to return in rat hindlimb muscles. Here we investigate how this recovery is influenced by experimental conditions. In an initial operation, the nerve of gastrocnemius medialis (GM) of adult rats was cut close to the muscle and reinserted either (i) close to the original nerve entry, or (ii) at a more medial 'foreign' site. In other groups of animals, these nerve operations were combined with a rotation of the GM muscle around its longitudinal axis, trying to ascertain whether the position of the muscle within the limb was of importance for the reinnervation processes. In a control group the muscle was rotated but innervation remained intact. After 21 weeks, the GM muscles were removed from both hindlimbs. Cross-sections were cut at seven different levels along each muscle, and 'slow' type I fibers were identified after staining for myofibrillar ATPase. The topographical positions were mapped out for all type I fibers. In all reinnervated muscles, an extensive type I fiber grouping was seen, indicating a widespread respecification of muscle fiber properties by ingrowing 'slow' axons. Normal topographical directions of type I fiber regionalization were about equally well restored in groups with the nerve inserted at the original or at the foreign site. In rotated muscles, the direction of type I fiber regionalization was significantly less rotated than the muscle as a whole. The results suggest that ingrowing 'slow' motor axons are guided toward their normal 'slow' regions by clues which are largely independent of the i.m. path of regeneration (original vs. foreign nerve entry site) but partly dependent on the position of the target muscle within the limb (rotated vs. non-rotated cases). (C) 2002 IBRO. Published by Elsevier Science Ltd. All rights reserved.

The topographical distribution of different fibre types in muscles of the lower hindlimb in rabbits and mice was quantitatively determined. The results were compared to those previously obtained, using the same new quantification methods, in homologous muscles of the rat. Type I fibres ('slow') were identified using myofibrillar ATPase histochemistry and mapped out at the mid proximo-distal level for 11 'fast' muscles in the rabbit and 7 'fast' muscles in the mouse. For the slow soleus muscle the procedure was undertaken for the type II fibres. Furthermore, for 5 of the 'fast' muscles from each animal species (extensor digitorum longus; flexor digitorum and hallucis longus; gastrocnemius medialis; peroneus longus; tibialis anterior), several more proximal and distal cross-sectional levels were also analysed. All the investigated 'fast' muscles showed a significant degree of topographical eccentricity in the midlevel distribution of type I fibres. For most muscles, the direction of this 'vector regionalisation' of type I fibres was similar between the three animal species. For homologous muscles, the degree of vector regionalisation was significantly different: mouse > rat > rabbit. The relative area of the region containing the type I fibres, inversely related to the degree of 'area regionalisation', was also significantly different: mouse

Newly developed concepts and methods for the quantification of fibre type regionalisation were used for comparison between all muscles traversing the ankle of the rat lower hindlimb (n = 13). For each muscle, cross-sections from the proximodistal midlevel were stained for myofibrillar ATPase and classified as type I (' slow ') or II (' fast '). For the 11 ' fast ' muscles (i.e. all except soleus), the muscle outline and the position of each type I fibre were digitised for further computer processing. Two potentially independent aspects of type I fibre regionalisation were evaluated quantitatively: (1) the degree to which type 1 fibres were restricted to a limited portion of the total cross-sectional area (' area-regionalisation '); (2) the extent and direction of the difference (if any) between the centre of the muscle cross-section and the calculated centre for the type I fibre cluster (' vector regionalisation '). Statistical analysis showed that type I fibres were vector regionalised in practically all investigated muscles and area regionalised within most of them, the only consistent exceptions being peroneus brevis and peroneus digitorum 4, 5. In muscles with a high degree of area regionalisation the population of type I fibres also had a markedly eccentric intramuscular position (i.e. high vector regionalisation). A significant relationship was observed between the relative position of a muscle within the hindlimb (transverse plane) and the direction and degree of its type I fibre eccentricity. On average, the degree of type I fibre eccentricity was greater for muscles remote from the limb centre than for those situated more centrally. In addition, the intramuscular concentration of type I fibres was typically greatest towards the centre of the limb, the most striking exception being tibialis posterior. For the slow soleus muscle, which is centrally placed within the limb, our analysis concerned the type II fibres, which were found to be weakly vector regionalised but not significantly area regionalised. It is concluded that, within muscles of the rat's lower hindlimb, fibre type regionalisation is a general and graded phenomenon which may reflect differentiating (embryological?) mechanisms of a transmuscular significance. Furthermore, the analysis demonstrated the usefulness of our new methods and concepts for the quantification of fibre type regionalisation.

Five muscles of the rat's lower hindlimb were compared with regard to their histochemical fibre type distribution at seven different proximo-distal levels. The muscles were: extensor digitorum longus (ED), flexor digitorum and hallucis longus (FD), gastrocnemius medialis (GM), peroneus longus (PE) and tibialis anterior (TA). In all the five muscles, the relative density of the 'slow' type I fibres showed a striking and similar decrease from proximal toward more distal levels. In addition, the type I fibres were concentrated within smaller and more eccentrically placed regions at distal than at more proximal levels. As a background for the further analysis of these lengthwise aspects of type I fibre regionalization, architectural features of the muscles were determined. Pinnation angles and the position of major tendons and tendon sheets were assessed in fresh specimens. Muscle fibre lengths were measured for single fibres dissected from different regions of macerated muscles. In all cases, fibre length was much shorter than muscle length (mean fraction ranging from 21 to 55%), implying that the proximo-distal changes in histochemical fibre properties were indeed explainable as being due to gradual lengthwise changes in fibre type populations. The similarity of these lengthwise changes across the muscles was in contrast to their differences in other aspects of functional organization, such as the average density of type I fibres and architectural features determining their relative capacities for shortening and force generation. The possible functional role of the proximal accumulation of type I fibres was discussed in relation to circulation and thermal balance; besides, the proximo-distal regionalization of type I fibres might (partly) reflect processes that had been associated with early stages of muscle differentiation. Furthermore, the results underline that, when determining the fibre type composition of rat hindlimb muscles, identifying the proximo-distal level of sampling is a matter of great importance.

To study how retinal ganglion cell axons diverge in the optic chiasm, the behavior of dye-labeled fibers was monitored in real time with video microscopy in an isolated preparation of embryonic mouse brain, with a focus on embryonic day 15-16. These real-time studies have revealed the dynamics of the growth of individual retinal axons, especially the tempo of extension and growth cone behaviors during divergence in the chiasm, a model for "decision" regions in developing pathways. Within the chiasm, retinal growth cones extend by saltatory growth, consisting of bursts of rapid advance alternating with pauses in extension. During pauses, growth cone appendages remain motile, and develop asymmetries prior to a change in the axis of growth. In a zone straddling the midline, retinal fibers, irrespective of destination, display long pauses for up to several hours, making small advances and retractions with no net extension. While crossed fibers ultimately progress through the midline, uncrossed fibers from inferior temporal retina develop wide-ranging branched growth cones, and then turn back to the ipsilateral side. Turns are effected by the selective retraction or micropruning of asymmetric foci of motile activity, and by the transformation of a backward-directed filopodium into a new growth cone. The behavior of retinal axons at the midline supports the hypothesis that this locus contains cues important for retinal axon divergence. Moreover, the observations of growth cone kinetics in the chiasm elucidate which growth cone forms seen in static preparations mediate growth cone turning, and suggest a model of axon navigation in decision regions in the intact nervous system.

International audience; Germline RET mutations are responsible for different inherited disorders: Hirschsprung disease (congenital aganglionic megacolon), caused by loss of function mutations, familial medullary thyroid carcinoma and multiple endocrine neoplasia type 2, caused by gain of function mutations. Intriguingly, some RET mutations, including C620R, are associated with both types of diseases. To investigate the dual role of such RET mutations, a mouse model with a targeted mutation ret(C620R) was generated. ret(C620R/C620R) offspring die during the first postnatal day, and show kidney agenesis and intestinal aganglionosis. Decreased outgrowth of the Ret-positive cells was observed in ret(C620R/C620R) neuronal cell cultures, which is suggestive of an impaired migration, proliferation or survival of the Ret-expressing cells. Electronmicroscopy revealed the absence of membrane-bound Ret in ret(C620R/C620R) cells as compared to ret(+/+) and ret(+/C620R) cells. On the other hand, aged ret(+/C620R) mice develop precancerous lesions in the adrenal gland or in the thyroid. Our results suggest that the ret(C620R) mutation has a loss of function effect in homozygotes and exhibits a dominant gain of function effect with low penetrance causing hyperplasia in heterozygotes.