This review leads us to a number of conclusions and suggestions for further study. First, we find wide differences in the meaning of flow heterogeneity, arising as a result of the different methods used. These differences will have to be reconciled to form a comprehensive view of the role of heterogeneity in determining vascular function. Second, in the future, the meaning of heterogeneity must be clearly defined and related to a particular microvascular component, and it is imperative that the differences in scale of heterogeneity be appreciated when comparing data from various laboratories. These heterogeneities have different implications for function, and failure to distinguish among them leads to confusion. Third, the degree to which perfusion heterogeneity is regulated in the microcirculation remains in doubt. Reports of variations in flow heterogeneity in response to physiological stimuli are for the most part based on highly questionable indirect methods. Fourth, the heterogeneity that can be demonstrated at the capillary level within striated muscle does not appear to be large relative to the capacity for the microcirculation to exchange most diffusible solutes. Thus, the inferences regarding heterogeneity, as evidenced by diffusible indicators, are likely to be the result of different preparations, damage to the preparations, or perhaps large-scale heterogeneities in the tissue. An alternate possibility would be that the heterogeneity occurs at the microvascular level but reflects some other aspect of microcirculatory function, such as length or hematocrit heterogeneities, but not flow heterogeneities. Fifth, flow heterogeneity within microvessels implies important consequences for capillary exchange and tissue oxygenation. Heterogeneities of velocity of a magnitude comparable to those observed by direct visualization of microcirculation can clearly produce reductions in oxygen supply to small tissue regions of a degree that may limit oxygen delivery, and thereby, tissue function. Sixth, flow heterogeneity may also influence capillary hematocrit and/or red cell spacing by producing cell separation at bifurcations and a resultant reduction in mean capillary tube hematocrit. There is as yet no agreement on why and how these hematocrits influence tissue oxygenation and function. Although several hypotheses are advanced to explain the distribution of blood flow and red cells within microcirculation, each lacks a critical experimental test at present.(ABSTRACT TRUNCATED AT 400 WORDS)

Abstract. Nerve growth factor (NGF), acidic fibroblast growth factor (aFGF), and basic fibroblast growth factor (bFGF) promote the survival and differentiation of a variety of peripheral and central neurons. The signal transduction mechanisms that mediate the actions of these factors in neuronal cells are not well understood. We examined the effect of a deficiency in protein kinase C (PKC) and/or cAMP second messenger systems on the actions of NGF, aFGF, and bFGF in the pheoehromocytoma (PC12) cell line. Activation of PKC was not required for NGF, aFGF, and bFGF to maximally induce ornithine decarboxylase (ODC), transcription of the early response genes, d2 and d5, or neurite outgrowth. In a PC12 cell mutant that is deficient in cAMP responsiveness (A126-1B2), all three growth factors maximally induced the transcription of d5 and neurite outgrowth, but aFGF and bFGF did not induce significant increases in ODC. NGF and aFGF maximally induced the transcription of d2 in A126-1B2 cells, but bFGF-induced d2 transcription was attenuated. NGF, aFGF, and bFGF maximally induced neurite outgrowth and d5 transcription in A126 cells that were made deficient in PKC. The d2 transcriptional response was substantially reduced in cells deficient in both PKC and cAMP responsiveness. These observations lead us to conclude that (a) cAMPand PKC-dependent events are, at least in part, causally linked to NGF, aFGF, and bFGF induction of both ODC and transcription of d2 and may control functionally redundant pathways; (b) NGF, aFGF, and bFGF can elicit neurite outgrowth and increase transcription of d2 and d5 in PC12 cells via mechanisms that are independent of both PKC and cAMP; (c) NGF, aFGF, and bFGF can induce ODC in the absence of PKC; and (d) aFGF and bFGF require cAMP responsiveness to induce ODC in PC12 cells.

The mechanism(s) by which heparin influences the biological activities of acidic and basic fibroblast growth factors (aFGF and bFGF) is not completely understood. One mechanism by which heparin could alter the biological activities of aFGF and bFGF is by altering their biological half-lives. We investigated the possibility that heparin potentiates aFGF-induced neurite outgrowth from PC12 cells by prolonging its biological half-life. Under conditions where heparin potentiated aFGF-induced neurite outgrowth, we observed that heparin increased the biological half-life of aFGF from 7 to 39 hr. We determined that greater than 25 hr of exposure to active aFGF was required for induction of neurite outgrowth. If aFGF activity was maintained for greater than 25 hr by periodic readdition of factor, heparin no longer potentiated aFGF-induced neurite outgrowth. These observations strongly suggest that heparin potentiates the activity of aFGF by prolonging its biological half-life. The protease inhibitors hirudin, leupeptin, and pepstatin A did not potentiate aFGF-induced neurite outgrowth, indicating that proteases inhibited by these inhibitors are not responsible for the loss of aFGF activity that we observed. However, aprotinin potentiated aFGF neurite-promoting activity approximately sevenfold, indicating that proteases that are inhibited by aprotinin are at least partially responsible for aFGF inactivation. These observations suggest that heparin regulates the activity of aFGF by regulating its proteolytic degradation, thereby regulating its biological half-life.

The sympathetic nervous system, via postganglionic innervation of blood vessels and the heart, is an important determinant of cardiovascular function. The mechanisms underlying sympathetic innervation of targets are not fully understood. This study tests the hypothesis that target-derived vascular endothelial growth factor (VEGF) promotes sympathetic innervation of blood vessels. Western blot and immunohistochemical analyses indicate that VEGF is produced by vascular cells in arteries and that VEGF receptors are expressed on sympathetic nerve fibers innervating arteries. In vitro, exogenously added VEGF and VEGF produced by vascular smooth muscle cells (VSMCs) in sympathetic neurovascular cocultures inhibited semaphorin 3A (Sema3A)-induced collapse of sympathetic growth cones. In the absence of Sema3A, VEGF and VSMCs also increased growth cone area. These effects were mediated via VEGF receptor 1. In vivo, the neutralization of VEGF inhibited the reinnervation of denervated femoral arteries. These data demonstrate that target-derived VEGF plays a previously unrecognized role in promoting the growth of sympathetic axons.