The accumulation of lysosomes and their hydrolases within neurons is a well-established neuropathologic feature of Alzheimer disease (AD). Here we show that lysosomal pathology in AD brain involves extensive alterations of macroautophagy, an inducible pathway for the turnover of intracellular constituents, including organelles. Using immunogold labeling with compartmental markers and electron microscopy on neocortical biopsies from AD brain, we unequivocally identified autophagosomes and other prelysosomal autophagic vacuoles (AVs), which were morphologically and biochemically similar to AVs highly purified from mouse liver. AVs were uncommon in brains devoid of AD pathology but were abundant in AD brains particularly, within neuritic processes, including synaptic terminals. In dystrophic neurites, autophagosomes, multivesicular bodies, multilamellar bodies, and cathepsin-containing autophagolysosomes were the predominant organelles and accumulated in large numbers. These compartments were distinguishable from lysosomes and lysosomal dense bodies, previously shown also to be abundant in dystrophic neurites. Autophagy was evident in the perikarya of affected neurons, particularly in those with neurofibrillary pathology where it was associated with a relative depletion of mitochondria and other organelles. These observations provide the first evidence that macroautophagy is extensively involved in the neurodegenerative/regenerative process in AD. The striking accumulations of immature AV forms in dystrophic neurites suggest that the transport of AVs and their maturation to lysosomes may be impaired, thereby impeding the suspected neuroprotective functions of autophagy.

Macroautophagy, a major pathway for organelle and protein turnover, has been implicated in the neurodegeneration of Alzheimer's disease (AD). The basis for the profuse accumulation of autophagic vacuoles (AVs) in affected neurons of the AD brain, however, is unknown. In this study, we show that constitutive macroautophagy in primary cortical neurons is highly efficient, because newly formed autophagosomes are rapidly cleared by fusion with lysosomes, accounting for their scarcity in the healthy brain. Even after macroautophagy is strongly induced by suppressing mTOR (mammalian target of rapamycin) kinase activity with rapamycin or nutrient deprivation, active cathepsin-positive autolysosomes rather than LC3-II-positive autophagosomes predominate, implying efficient autophagosome clearance in healthy neurons. In contrast, selectively impeding late steps in macroautophagy by inhibiting cathepsin-mediated proteolysis within autolysosomes with cysteine-and aspartyl-protease inhibitors caused a marked accumulation of electron-dense double-membrane-limited AVs, containing cathepsin D and incompletely degraded LC3-II in perikarya and neurites. Similar structures accumulated in large numbers when fusion of autophagosomes with lysosomes was slowed by disrupting their transport on microtubules with vinblastine. Finally, we find that the autophagic vacuoles accumulating after protease inhibition or prolonged vinblastine treatment strongly resembled AVs that collect in dystrophic neurites in the AD brain and in an AD mouse model. We conclude that macroautophagy is constitutively active and highly efficient in healthy neurons and that the autophagic pathology observed in AD most likely arises from impaired clearance of AVs rather than strong autophagy induction alone. Therapeutic modulation of autophagy in AD may, therefore, require targeting late steps in the autophagic pathway.

A defect in the structure of the obese gene is responsible for development of obesity in the ob͞ob mouse. The product of expression of the gene is the protein hormone leptin. Leptin causes weight loss in ob͞ob and normal mice, it is secreted by adipocytes, and it is an important controller of the size of fat stores by inhibiting appetite. The ob͞ob mouse is infertile and has a pattern of gonadotropin secretion similar to that of prepubertal animals. Consequently, we hypothesized that leptin might play a role in the control of gonadotropin secretion and initiated studies on its possible acute effects on hypothalamic-pituitary function. After a preincubation period, hemi-anterior pituitaries of adult male rats were incubated with leptin for 3 hr. Leptin produced a dose-related increase in follicle-stimulating hormone (FSH) and luteinizing hormone (LH) release, which reached peaks with 10 ؊9 and 10 ؊11 M leptin, respectively. Gonadotropin release decreased at higher concentrations of leptin to values indistinguishable from that of control pituitaries. On the other hand, prolactin secretion was greatly increased in a dose-related manner but only with leptin concentrations (10 ؊7 -10 ؊5 M). Incubation with leptin of median eminence-arcuate nuclear explants from the same animals produced significant increases in LH-releasing hormone (LHRH) release only at the lowest concentrations tested (10 ؊12 -10 ؊10 M). As the leptin concentration was increased, LHRH release decreased and was significantly less than control release at the highest concentration tested (10 ؊6 M). To determine if leptin can also release gonadotropins in vivo, ovariectomized females bearing implanted third ventricle cannulae were injected with 10 g of estradiol benzoate s.c., followed 72 hr later by microinjection into the third ventricle of leptin (0.6 nmol in 5 l) or an equal volume of diluent. There was a highly significant increase in plasma LH, which peaked 10-50 min after injection of leptin. Leptin had no effect on plasma FSH concentrations, and the diluent had no effect on either plasma FSH or LH. Thus, leptin at very low concentrations stimulated LHRH release from hypothalamic explants and FSH and LH release from anterior pituitaries of adult male rats in vitro and released LH, but not FSH, in vivo. The results indicate that leptin plays an important role in controlling gonadotropin secretion by stimulatory hypothalamic and pituitary actions.