Ghrelin, an endogenous growth hormone secretagogue, is a novel orexigenic peptide that antagonizes leptin action through the activation of hypothalamic neuropeptide Y/Y1 receptor pathway.

Ghrelin, an endogenous ligand for growth hormone secretagogue (GHS) receptor originally isolated from the stomach, occurs in the hypothalamic arcuate nucleus and may play a role in energy homeostasis. Synthetic GHSs have activated the hypothalamic arcuate neurons containing neuropeptide Y (NPY), suggesting the involvement of NPY in some of ghrelin actions. This study was designed to elucidate the role of ghrelin in the regulation of food intake. A single intracerebroventricular (ICV) injection of ghrelin (5-5,000 ng/rat) caused a significant and dose-related increase in cumulative food intake in rats. Ghrelin (500 ng/rat) was also effective in growth hormone-deficient spontaneous dwarf rats. Hypothalamic NPY mRNA expression was increased in rats that received a single ICV injection of ghrelin (500 ng/rat) (approximately 160% of that in vehicle-treated groups, P < 0.05). The ghrelin's orexigenic effect was abolished dose-dependently by ICV co-injection of NPY Y1 receptor antagonist (10-30 microg/rat). The leptin-induced inhibition of food intake was reversed by ICV co-injection of ghrelin in a dose-dependent manner (5-500 ng/rat). Leptin reduced hypothalamic NPY mRNA expression by 35% (P < 0.05), which was abolished by ICV co-injection of ghrelin (500 ng/rat). This study provides evidence that ghrelin is an orexigenic peptide that antagonizes leptin action through the activation of hypothalamic NPY/Y1 receptor pathway.

Electron microscopy of unstained, fresh air-dried spreads of the mouse brain and application to energy dispersive X-ray microanalysis.

Electron microscopy of unstained, fresh air-dried spreads of the mouse brain revealed well-preserved ultrastructures of neurons and synaptic boutons filled with synaptic vesicles. Energy dispersive X-ray microanalysis of the neuron nucleus disclosed peaks for phosphorus, sulfur, chlorine and potassium. Analysis of large regions (spots of ca. 1 micron in diameter) on the neuron nucleus revealed peaks for magnesium, whereas that of regions on the cytoplasm rarely disclosed the element, which was confirmed by removal of the background noise using a micro-Edit computer system. Analysis of regions on synaptic boutons except mitochondria also indicated peaks for magnesium.

Eelctron microscopy of unstained fresh air-dried spreads of mouse pancreas acinar cells and energy dispersive x-ray microanalysis of zymogen granules.

Electron microscopy of unstained, fresh air-dried spreads of the mouse pancreas has disclosed various substructures of acinar cells. The nucleus with a nucleolus, rough endoplasmic reticulum with ribosomes, mitochondria with dense granules, lysosomes, intracisternal granules, and zymogen granules with a core surrounded by a halo, and a boundary membrane were observable under a conventional electron microscope. Five zymogen granules of an acinar cell were examined by energy dispersive X-ray microanalysis. Peaks for phosphorus, sulfur, chlorine and potassium were detected in all of the zymogen granules examined, which confirmed a previous report (Takaya, 1975b). By increasing the integrated pulse counting time up to 300 seconds, calcium and magnesium were detected in two of the granules in addition to the elements mentioned above. No peaks for either copper or zinc were observed when nylon grids were used. The electrolyte content of zymogen granules and of acinar initial fluid is discussed in relation to the secretory mechanism of the exocrine pancreas.

The variable refractivity of the protein or polypeptide hormone-producing cells showing a unique luminescence in the dark-field microscope.

When cryostat sections of endocrine tissue were examined in a dark-field microscope, a brillant granular luminescence was revealed in the endocrine cells thought to be concerned with protein or polypeptide horome production. The sections were prepared from fresh materials either frozen in a cryostat chamber at --25 degrees C, in dry ice-acetone, or fixed in formalin-calcium for 24 hr. The neurosecretory substance in the hypothalamus and the posterior lobe of the pituitary showed a blue luminescence; the acidophil cells of the anterior lobe of the pituitary, orange; basophile cell, green or blue; intermediate lobe cells, no luminescence; thyroid C cells, white-blue; pancreatic A cells, blue; B cells, orange; adrenomedullary cells, greenish blue; enterochromatin cells, green; and other endocrine cells in the gastrointestinal tract, blue or orange. After tearing and spreading the pituitary and hypothalamus with a pair of needles on a glass slide, and examining the teased specimen by dark-field microscopy, various cells of different lumunescent colours became apparent in the anterior lobe of the pituitary, a blue fluorescent substance in the posterior lobe, and neurosecretory cells bodies in the hypothalamus. The different colours appear to be inherent in the granules of living tissues.

Energy dispersive x-ray microanalysis of neurosecretory granules of mouse pituitary on fresh air-dried tissue spreads.

Electron microscopy of fresh air dried spreads of unstained posterior lobe tissue from mouse pituitary disclosed neurosecretory granules. Each granule showed a seemingly homogeneous dense core surrounded by a halo and a bounding membrane. The area between granules in the cytoplasm was relatively well preserved. The energy dispersive X-ray microanalysis revealed peaks for sulfur, chlorine and potassium in two granules. The third granule displayed peaks for phosphorus and chlorine. These elements probably contribute to the high electron density of the granules. There was no peak for calcium, in contrast to the dense bodies of human blood platelets.