GlcNAc6ST-1 regulates sulfation of N-glycans and myelination in the peripheral nervous system.

Highly specialized glial cells wrap axons with a multilayered myelin membrane in vertebrates. Myelin serves essential roles in the functioning of the nervous system. Axonal degeneration is the major cause of permanent neurological disability in primary myelin diseases. Many glycoproteins have been identified in myelin, and a lack of one myelin glycoprotein results in abnormal myelin structures in many cases. However, the roles of glycans on myelin glycoproteins remain poorly understood. Here, we report that sulfated N-glycans are involved in peripheral nervous system (PNS) myelination. PNS myelin glycoproteins contain highly abundant sulfated N-glycans. Major sulfated N-glycans were identified in both porcine and mouse PNS myelin, demonstrating that the 6-O-sulfation of N-acetylglucosamine (GlcNAc-6-O-sulfation) is highly conserved in PNS myelin between these species. P0 protein, the most abundant glycoprotein in PNS myelin and mutations in which at the glycosylation site cause Charcot-Marie-Tooth neuropathy, has abundant GlcNAc-6-O-sulfated N-glycans. Mice deficient in N-acetylglucosamine-6-O-sulfotransferase-1 (GlcNAc6ST-1) failed to synthesize sulfated N-glycans and exhibited abnormal myelination and axonal degeneration in the PNS. Taken together, this study demonstrates that GlcNAc6ST-1 modulates PNS myelination and myelinated axonal survival through the GlcNAc-6-O-sulfation of N-glycans on glycoproteins. These findings may provide novel insights into the pathogenesis of peripheral neuropathy.

Detection of N-glycans on small amounts of glycoproteins in tissue samples and sodium dodecyl sulfate-polyacrylamide gels.

N-linked glycans harbored on glycoproteins profoundly affect the character of proteins by altering their structure or capacity to bind to other molecules. Specific knowledge of the role of N-glycans in these changes is limited due to difficulties in identifying precise carbohydrate structures on a given glycoprotein, which arises from the large amounts of glycoprotein required for N-glycan structural determination. Here, we refined a simple method to purify and detect trace amounts of N-glycans. During the N-glycan purification step, most contaminants were removed by two kinds of columns: a graphite carbon column and a cellulose column. N-Glycans were identified with a three-dimensional high-performance liquid chromatography (HPLC) system. Using our method, a global analysis of N-glycans from human muscle biopsy samples and mouse brain sections was possible. By combining sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with our method, we refined analytical procedures for N-glycans from SDS-PAGE gels using hydrazinolysis to achieve a high N-glycan recovery rate. N-Glycans on as little as 1 µg of the target protein transferrin or immunoglobulin G (IgG) were easily detected. These methods allowed us to efficiently determine glycoprotein N-glycans at picomole (pmol) levels.

Evaluation of a medetomidine-midazolam combination for immobilizing and sedating Japanese monkeys (Macaca fuscata).

We clinically and clinicopathologically investigated the immobilizing and sedative effects of a medetomidine-midazolam (MM) combination in Japanese monkeys (Macaca fuscata) and its antagonism with atipamezole. MM (medetomidine, 60 microg/kg; midazolam, 0.3 mg/kg) was administered intramuscularly to each monkey (n = 11). All animals were laterally recumbent within 13 +/-6 min after administration of MM. This combination induced deep sedation accompanied by analgesia, muscle relaxation, and markedly depressed arousal reactions to external stimuli. After administration of atipamezole (240 microg/kg intramuscularly), the animals recovered rapidly and smoothly to their normal postures within 10 +/-2 min. In this study, the hematologic and serum biochemical parameters of Japanese monkeys given MM did not differ significantly from those of Japanese monkeys under general anesthesia via ketamine. Salivary a-amylase activities (stress indexes) ranged from 4 to 99 kU/l in Japanese monkeys, similar to levels measured in humans. An important advantage of MM was that its effects were reversible with atipamezole. We have confirmed that MM is valuable as a chemical restraint agent in Japanese monkeys for various experimental procedures.

Stroop performance and difficulties of older adults using automatic teller machines.

This study examined the relationship between inhibition of older dults and difficulties using automatic teller machines. 22 older adults performed three transactions on an ATM simulator, and errors were recorded. Participants also performed the Stroop interference task to assess inhibition. Both interference effect in response latencies and rate of intrusion errors in the Stroop task were moderately correlated with number of errors on the task. Older adults with low Stroop performance made more errors in the categories of Forgetting an operation, Screen mix-up, and Overlooking an input error than those with high performance. These results suggest that low inhibition may be associated with difficulties in automatic teller machine use, and the importance of designing a user interface which lessens the likelihood of errors for older adults or others.