Development of cerebellar astroglia: Transitions in form and cytoskeletal content

The forms, disposition, and cytoskeletal contents of astroglia in immature mouse cerebellum were studied by immunocytochemical staining with antisera against two intermediate filament proteins, vimentin (Vim) (58,000 daltons) and glial filament protein (GF) (51,000 daltons). From embryonic (E) Day 15 to postnatal (P) Day 2, Vim is expressed in cells throughout the cerebellar anlage, including radial glia and Bergmann fibers, cells with amorphous shapes and 2–3 processes, and thick longitudinal elements oriented parallel to axons within axon tracts. GF is not expressed during the first few postnatal days, but by P7, there is a dramatic increase in GF-positive astrocyte-like cells in the putative white matter that are more densely stained and more crowded than at any other age. Between P7 and P14 all astrocytes throughout the cerebellum express both Vim and GF. From P21 on, Vim expression is progressively rarer in all astrocytes except for Bergmann fibers, and GF-positive astrocytes become less numerous. These findings raise two issues: (a) the lineage and relationships of cells expressing Vim and GF; (b) Since GF-positive cells appear as axon ingrowth ceases, axons must grow in a terrain comprised of glial cells that have a different cytoskeletal composition (vimentin), reflecting a less differentiated state, than mature astrocytes or than the GF-rich astrocytes that proliferate after injury in adult CNS.     

Molecular cloning of the Escherichia coli K-12 entACGBE genes

Abstract A 7-kb piece of Escherichia coli DNA that contains five genes ( entA, C, G, B and E ) required for the biosynthesis of the iron transport molecule enterochelin was isolated. A restriction map was constructed and proteins specified by the E. coli DNA were identified in mini- and maxicell systems. Plasmids containing portions of the entACGBE DNA generated by BAL31 digestion or restriction enzyme treatment were constructed; complementation studies done with these indicated that the five genes constitute an operon. The approximate site of the promoter was determined and the product of entE was tentatively identified as an M _r 63000 polypeptide.     

Localization of glyoxylic acid-induced histofluorescence in surgically isolated medial basal hypothalamus of the rat

Summary Examination of glyoxylic acid-induced catecholamine histofluorescence in the hypothalamic median eminence of adult male rats revealed a linear pattern of fine varicosities coursing through the ependymal and fibrous zones, suggestive of juxtaposition to tanycytes. In order to determine the origin of these terminals, adult rats were subjected to complete isolation of the medial basal hypothalamus, using a small Halasz-Pupp knife. As rapidly as 24h after this deafferentation degenerative axon profiles were observed dorsal, as well as anterior and lateral, to the knife track. Occasionally at three days postoperatively, and routinely by seven days after surgery, fine-sized new fibres were seen passing through the knife wound. The linear profiles of varicosities observed in the normal median eminence remained traceable in the experimental preparations; the site of origin for these terminals therefore appears to be neurons of the arcuate (A12) and rostral periventricular (A14) regions. The results also indicate that fibres innervating the isolated area are capable of morphologically demonstrable new growth. The observations bear functional implications in assessing endocrine regulation following MBH isolation of the type used in this study.This study was supported by USPHS Postdoctoral Fellowship 5-F₂2-HD00630 (CT) and USPHS Grant NS-11642 (JRS). The authors wish to express their appreciation to Yvonne Cheung and Patricia Walker for technical assistance     

Inhibition of thymidine uptake in asynchronous HeLa cells by nitrobenzylthioinosine

Nitrobenzylthioinosine (NBMPR), an inhibitor of nucleoside transport by human erythrocytes, was found to be a potent inhibitor of thymidine uptake by asynchronous monolayer cultures of HeLa cells. Rates of thymidine uptake by the cultures at 20 °C were constant between 10 and 40 sec after thymidine addition and were assayed during this interval; TTP was the principal metabolite of thymidine and the thymidine phosphates accumulated at constant rates which extrapolated through time zero. The lack of an effect of NBMPR on thymidine kinase activity, or on the relative proportions of thymidine metabolites in cell extracts, indicated that NBMPR inhibited thymidine transport. When mediated entry (transport) was eliminated by 2 μM NBMPR, a significant diffusional component of thymidine entry was apparent. The mediated component of thymidine uptake exhibited Michaelis-Menten kinetics and apparent K_m and V_max values of 0.5 μM and 10–21 pmoles/min/10⁶ cells were obtained. When NBMPR-treated cells were transferred to NBMPR-free medium, inhibition of thymidine uptake persisted, suggesting that NBMPR was firmly bound to the transport inhibitory sites.     

Sexual preferences of male hamsters (Mesocricetus auratus) for conspecifics in different endocrine conditions

The behavioral responses of sexually experienced male hamsters toward a pair of anesthetized conspecifics were investigated. Males spent significantly more time licking, sniffing, and mounting neonatally and adult castrated males than intact males. Adult castrated males receiving oil injections were preferred over castrates receiving exogenous testosterone propionate (TP). Ovariectomized females were preferred over intact males, adult castrated males, or spayed females receiving exogenous TP. It was concluded that the absence of an androgen-dependent factor(s) renders an animal more sexually attractive.     

Abnormal functional development of sympathetic nerve terminal-smooth muscle complex in neonatal spontaneously hypertensive rats

In neonatal and young adult SH and WK rats, maturation of a functional unit consisting of sympathetic nerve terminals and smooth muscle (levator palpebrae) was assessed $\text{in vivo}$ by measuring the contractile response to tyramine, an agent which releases endogenous norepinephrine from sympathetic nerve terminals. Responses in SH are comparable to WK at 5–6 days postnatally, smaller from the 8th through 19th day and larger in young adults (41–46 days old). Results indicate that functional maturation of the nerve terminal-smooth muscle complex is retarded in SH relative to WK during the 2nd and 3rd postnatal weeks. It is suggested that retardation in the neonatal SH rat is an expression of a genetic defect in the growth of the complex and that the enhanced response in young adult SH is a consequence of the neonatal abnormality.     

Ionic charge on phospholipids and their interaction with the mitochondrial adenosine triphosphatase

The activity of the lipid-depleted, oligomycin-sensitive mitochondrial ATPase has been measured in the presence of liposomes prepared from mixtures of phosphatidylglycerol and phosphatidylglycerol lysine. Enzyme activity increased linearly with an increase in the negative charge of liposomes prepared from the phosphatidylglycerol-phosphatidylglycerol lysine mixtures. The electrophoretic mobility and activating capacity of liposomes of several other phospholipids were determined. A linear relationship between electrophoretic mobility of the liposomes and oligomycin-sensitive activity was again apparent. These observations demonstrate that the activity of the ATPase is directly proportional to the ionic charge on phospholipid activators if the acyl chain composition of the phosphoglycerides is relatively constant.     

The immune response of inbred mouse strains to DNP-BGG

The immune response of six inbred mouse strains (SJL, A, C57BL/6, CBA, BALB/c, and DBA/1) to DNP₅₆BGG was tested under three separate immunization schedules: 1 Μg DNP-BGG in 1 mg Al(OH)₃ adjuvant, 50 Μg DNP-BGG in 1 mg A1(OH)₃ adjuvant, and 1 Μg DNP-BGG in complete Freund's adjuvant. Individual serum samples were titered using a modified Farr assay. It was found that the first schedule allowed classification of the mice into responder (SJL, A) and nonresponder (C57BL/6, CBA, BALB/ c, DBA/1) strains. The second schedule produced quantitative as well as qualitative differences among the strains and allowed classification of the mice into higher-responder (SJL, A), intermediate-responder (C57BL/6, CBA, BALB/c), and low-responder (DBA/1) categories. When complete Freund's adjuvant was used in the third schedule, the differences among strains became insignificant. The sera from each strain were pooled and assayed for relative antibody affinity and IgM content. Both of these parameters were dependent largely on the dose of antigen and type of adjuvant used, rather than on the particular mouse strain being studied. The mechanism of adjuvant action, and possible cell interactions in the genetic control of the immune response, are discussed.