Morphological classification of bovine ovarian follicles

Follicle classification is an important aid to the understanding of follicular development and atresia. Some bovine primordial follicles have the classical primordial shape, but ellipsoidal shaped follicles with some cuboidal granulosa cells at the poles are far more common. Preantral follicles have one of two basal lamina phenotypes, either a single aligned layer or one with additional layers. In antral follicles <5 mm diameter, half of the healthy follicles have columnar shaped basal granulosa cells and additional layers of basal lamina, which appear as loops in cross section (‘loopy’). The remainder have aligned single-layered follicular basal laminas with rounded basal cells, and contain better quality oocytes than the loopy/columnar follicles. In sizes >5 mm, only aligned/rounded phenotypes are present. Dominant and subordinate follicles can be identified by ultrasound and/or histological examination of pairs of ovaries. Atretic follicles <5 mm are either basal atretic or antral atretic, named on the basis of the location in the membrana granulosa where cells die first. Basal atretic follicles have considerable biological differences to antral atretic follicles. In follicles >5 mm, only antral atresia is observed. The concentrations of follicular fluid steroid hormones can be used to classify atresia and distinguish some of the different types of atresia; however, this method is unlikely to identify follicles early in atresia, and hence misclassify them as healthy. Other biochemical and histological methods can be used, but since cell death is a part of normal homoeostatis, deciding when a follicle has entered atresia remains somewhat subjective.

Morphine-induced receptor endocytosis in a novel knockin mouse reduces tolerance and dependence

Abstract Opioid drugs, such as morphine, are among the most effective analgesics available. However, their utility for the treatment of chronic pain is limited by side effects including tolerance and dependence. Morphine acts primarily through the mu-opioid receptor (MOP-R) , which is also a target of endogenous opioids. However, unlike endogenous ligands, morphine fails to promote substantial receptor endocytosis both in vitro, and in vivo. Receptor endocytosis serves at least two important functions in signal transduction. First, desensitization and endocytosis act as an "off" switch by uncoupling receptors from G protein. Second, endocytosis functions as an "on" switch, resensitizing receptors by recycling them to the plasma membrane. Thus, both the off and on function of the MOP-R are altered in response to morphine compared to endogenous ligands. To examine whether the low degree of endocytosis induced by morphine contributes to tolerance and dependence, we generated a knockin mouse that expresses a mutant MOP-R that undergoes morphine-induced endocytosis. Morphine remains an excellent antinociceptive agent in these mice. Importantly, these mice display substantially reduced antinociceptive tolerance and physical dependence. These data suggest that opioid drugs with a pharmacological profile similar to morphine but the ability to promote endocytosis could provide analgesia while having a reduced liability for promoting tolerance and dependence

Identifying optimal lipid raft characteristics required to promote nanoscale protein-protein interactions on the plasma membrane

The dynamic lateral segregation of signaling proteins into microdomains is proposed to facilitate signal transduction, but the constraints on microdomain size, mobility, and diffusion that might realize this function are undefined. Here we interrogate a stochastic spatial model of the plasma membrane to determine how microdomains affect protein dynamics. Taking lipid rafts as representative microdomains, we show that reduced protein mobility in rafts segregates dynamically partitioning proteins, but the equilibrium concentration is largely independent of raft size and mobility. Rafts weakly impede small-scale protein diffusion but more strongly impede long-range protein mobility. The long-range mobility of raft-partitioning and raft-excluded proteins, however, is reduced to a similar extent. Dynamic partitioning into rafts increases specific interprotein collision rates, but to maximize this critical, biologically relevant function, rafts must be small (diameter, 6 to 14 nm) and mobile. Intermolecular collisions can also be favored by the selective capture and exclusion of proteins by rafts, although this mechanism is generally less efficient than simple dynamic partitioning. Generalizing these results, we conclude that microdomains can readily operate as protein concentrators or isolators but there appear to be significant constraints on size and mobility if microdomains are also required to function as reaction chambers that facilitate nanoscale protein-protein interactions. These results may have significant implications for the many signaling cascades that are scaffolded or assembled in plasma membrane microdomains.