MATERIALS AND METHODS

The mice used in this study came from two separate sources: the normal control mice from the C57BL/KsJ inbred strain maintained by the production department of The Jackson Laboratory, and the diabetic db/db mice from the C57BL/KsHu-db strain maintained in our research colony. All mice were fed a commercial laboratory chow (Old Guilford Corp., Guilford, Conn.) containing 6% of fat. The diabetic mice are descendants of the original mutants and have been propagated by matings of C57BL/KsHu-db mice heterozygous at the db locus. In order to assess any histocompatibility differences which may have arisen at or since the time of the mutation, control parabiotic unions were made between normal mice of the C57BL/KsHu and the C57BL/KsJ strains. Parabiosis was not performed on weanling diabetic mice because of the delicate nature of both peritoneal wall and skin. Moreover, older diabetic mice become so grossly obese that parabiosis is technically difficult. Therefore, diabetic mice from 6 to 8 weeks of age that still had reasonably low blood sugar concentrations(usually< 250 mg/lOO ml) were placed on restricted intake of food for the period of time (25 to 93 days) necessary to reduce their weights to near normal (Table 1, group I). The dietary restriction consisted of feeding for 8 hr on Mondays, Wednesdays, and Fridays, only, while allowing access to water at all times. Blood sugar determinations(fasting levels) were made on Monday morning of each week prior to the first feeding period. Normal mice do not tolerate this severe schedule of food restriction and unrestricted normal mice were used in all experiments.

The technique of para biosis, carried out under sodium pentobarbital anesthesia (6.2 mg/lOO g), included celioanastomosis as well as anastomosis of the skin from the shoulder to the pelvic girdle. Silk sutures were used to join the peritoneal walls and a stay suture was placed through adjacent scapulas. Wound clips were used to effect skin union. After surgery, each pair of mice was housed separately and fed the laboratory chow ad libitum. Three types of parabiont pairs were produced; db/db with j-/-j--, db/db with db/db, and+/+ with+/+. Two groups of unparabiosed diabetic mice were maintained as controls; one that remained on the restricted feeding schedule throughout(restricted), and one that was fed ad libitum following a restricted feeding period (restricted refed). Weights were recorded weekly at the time that blood sugar concentrations were determined on the controls and on each animal in the parabiont pairs by the micromethod of Folin and Malmros on 50~ 1 of blood obtained from the orbital sinus (4). Plasma insulin was estimated by an