GRF-induced feeding: Evidence for protein selectivity and opiate involvement

Growth hormone-releasing factor (GRF) is a hypothalamic peptide named for its ability to induce release of growth hormone from the anterior pituitary. GRF also acts as a neurotransmitter in the suprachiasmatic nucleus/medial preoptic area (SCN/MPOA) to stimulate food intake. The purpose of this series of experiments was to explore the nature of GRF-induced feeding, with a particular emphasis on macronutrient selectivity, and to examine the role of opiate activity in the paraventricular nucleus of the hypothalamus (PVN). Chow intake stimulated by GRF microinjection (1 pmol/0.5 μl) into the SCN/MPOA was blocked by injection of methyl-naltrexone (3 μg/0.5 μl) into the PVN. In animals habituated to macronutrient diets (Teklad, WI), GRF preferentially stimulated intake of protein at 2 and 4 h postinjection, whereas it had no effect on carbohydrate intake. Further, this effect was blocked by injection of naloxone (40 nmol/0.5 μl) into the PVN. Microinjection of morphine (0, 1, 10, and 17 μg/0.5 μl) into the PVN also specifically stimulated protein intake at 2 and 4 h postinjection. These results suggest that feeding derived from GRF actions in the SCN/MPOA is macronutrient selective, and is dependent on PVN opiate activity for expression.     

Sperm cell surface characteristics of Plumbago zeylanica L. in relation to transport in the embryo sac

Myosin associated with the male germ cells of angiosperms interacts with actin, promoting transport of the non-motile generative and later sperm cells in the pollen tube. Myosin localizing on the sperm cell plasma membrane seems negligible in Plumbago, as reflected by the absence of: (i) anti-myosin labeling using immunoelectron microscopy, (ii) sperm motility on actin matrices, and (iii) electrophoretic movement changes after addition of antibody. Sperm cells injected directly into actively streaming Nitella internodal cells, however, follow actin bundles and their movement is sensitive to ATP and Mg²⁺. This may be based on simple charge binding since negatively charged latex beads also migrate on actin, whereas neutral or positively-charged latex beads do not. Sperm cells are negatively charged according to capillary microelectrophoresis, whereas killed sperm cells, which are positively charged do not migrate. The sperm cell that normally fertilizes the egg has a higher calculated charge (8.277 × 10³ esu/cm²) compared with the sperm cell that fuses with the central cell (6.120 × 10³ esu/cm²). Received: 15 December 1998 / Accepted: 21 January 1999     

Morphological correlates of serotonin-neuropeptide Y interactions in the rat suprachiasmatic nucleus: Combined radioautographic and immunocytochemical data

Summary The morphological substrate of putative serotonin (5-HT)/neuropeptide Y (NPY) interactions in thé suprachiasmatic nucleus (SCN) was investigated by combined radioautography and immunocytochemistry after intraventricular administration of (³H)5-HT in the rat. In the ventral portion of the SCN, the distribution of (³H)5-HT uptake sites overlapped closely the NPY-immunoreactive terminals. Previous investigations have shown that the dense 5-HT and NPY innervations of the SCN originate in different structures, i.e., the midbrain raphe nuclei and the ventral lateral geniculate nucleus, respectively. Accordingly, in the present study, destruction of 5-HT afferents by 5,7-dihydroxytryptamine was not found to induce any modification in NPY staining and, in ultrastructural immuno-radioautographic preparations, two distinct pools of axonal varicosities could be identified. Both 5-HT and NPY terminals established morphologically defined synaptic junctions, sometimes on the same neuronal target. Some cases of direct axo-axonic appositions between the two types of terminals were also encountered. These data constitute additional criteria for characterizing the cytological basis of the multiple transmitter interactions presumably involved in the function of the SCN as a central regulator of circadian biological rhythms.     

Localization of chitin on ultrathin sections of cysts of two ciliated protozoa,Blepharisma undulans andPseudomicrothorax dubius,using colloidal gold conjugated wheat germ agglutinin

Summary The cyst walls of the ciliatesBlepharisma undulans andPseudomicrothorax dubius were examined ultrastructurally and by postembedding labeling with wheat germ agglutinin (WGA)-gold conjugate. Different methods of fixation and embedding were performed. In all procedures, WGA-gold binds selectively to material of the cyst wall. Pretreatment of the sections with chitinase inhibits labeling. The cyst walls of both species contain 3 nm fibrils, which are supposed to be of chitinous nature. In the cyst wall ofB. undulans, several thin layers of WGA-binding fibrils are interspaced with thick layers of other material. InP. dubius, WGA-binding sites are mainly concentrated in the mesocyst, where the microfibrils appear to represent the major component. These results obtained from two phylogenetically distant species confirm that chitin synthesis is an ancestral feature of ciliated protozoa. The amount and distribution of the chitin fibrils may play an important role in the properties and functions of the wall of the resting cyst.     

Wheat germ agglutinin binding sites on human urothelial cells of different grades of transformation

¹²⁵I-Wheat germ agglutinin (WGA) binding parameters of human urothelial cell lines of different grades of transformation (TGrll and TGrlll) were compared. The values of association constant (K_a) and the number of binding sites/cell for HCV29 (TGrll) cell line were about 3×10⁶M^–1 and over 4×10⁷, respectively. Two TGrlll cell lines, HCV29T and Hu549 revealed lower values for K_a, and considerably higher numbers of binding sites/cell (about 3×10⁸ and 2×10⁸, respectively). Binding of¹²⁵I-WGA to total cellular proteins resolved by SDS-PAGE and transferred to nitrocellulose showed multiple diffused bands in the range of 58–180 kDa. Some of these bands were characteristic for TGrll cells (124 kDa) or TGrlll cells (135 and 148 kDa).Abbreviations TGr transformation grade - WGA wheat germ agglutinin - sWGA succinylated wheat germ agglutinin - GlcNAc N-acetyl-d-glucosamine - BSA bovine serum albumin - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis     

Catecholaminergic innervation of GRF-containing neurons in the rat hypothalamus revealed by electron-microscopic cytochemistry

Summary The Catecholaminergic innervation of neurons containing growth hormone-releasing factor (GRF) was examined by use of a method which combined either 5-hydroxydopamine (5-OHDA) uptake or autoradiography after intraventricular injection of ³H-noradrenaline with immunocytochemistry for GRF in the same tissue sections at the electron-microscopic level. In the ventrolateral part of the arcuate nucleus of the rat hypothalamus a large number of immunonegative axon terminals were found to make synaptic contact with GRF-like immunoreactive (GRF-LI) cell bodies and processes. ³H-noradrenaline autoradiography or 5-OHDA-labeling combined with GRF immunocytochemistry revealed that axon terminals labeled with ³H-noradrenaline or 5-OHDA make synaptic contact with the GRF-LI nerve cell bodies and processes. These findings indicate that catecholamine-containing neurons innervate GRF neurons to regulate GRF secretion via synapses in the rat arcuate nucleus.     

视前区微量注射吗啡对大鼠丘脑束旁核痛反应神经元电活动的影响

本实验观察了视前区(POA)内微量注入阿片样物质对丘脑束旁核(Pf)痛反应神经元电活动影响。结果如下:(1)POA 内微量注射高浓度吗啡(10μg/μl)能显著抑制 Pf 内大部分(20/26)痛兴奋神经元(PEN)的痛诱发放电,其中3个神经元注药后对伤害性刺激转变成抑制反应;POA 内微量注射低浓度吗啡(1μg/μl)也显著抑制 Pf 内大部分(19/23)PEN 的痛诱发放电。(2) POA 内微量注射两种浓度的吗啡,均使大多数痛抑制神经元(PIN,共27/33)的完全抑制时程缩短。上述结果提示,POA 内阿片样物质对 Pf 内痛反应神经元的电活动可能具有抑制作用。     

刺激大鼠下丘脑室旁核激活的中脑中央灰质神经元对躯体传入的反应

电刺激大鼠下丘脑室旁核(PVH),在同侧中脑中央灰质(CG)内寻找逆行及顺行反应单位,然后观察它们对躯体感觉刺激的反应。实验结果表明:CG 及邻近网状结构内有10%(32/318)的单位呈逆行反应。逆行传导速度平均为0.37±0.24m/s(均数±标准差);推测这种CG→PVH 投射纤维属于细有髓或无髓神经纤维。这些单位分布于 CG 的腹外侧及背外侧亚核。50%(14/28)的逆行单位对坐骨(胫)神经的强电刺激和夹尾等损伤性刺激起反应,但对触毛或低强度的神经干刺激无明显反应。以上结果表明:外周躯体感觉,特别是损伤性信息传入 PVH 时,CG 是其中枢驿站之一。电刺激 PVH 还能顺行激活7.55(24/318)、抑制0.7%(2/318)的 CG 单位。有69%(18/26)的顺行反应单位对外周躯体神经强电刺激及夹尾起反应。提示 PVH 可能通过影响 CG神经元的活动而参与中枢痛觉的整合。