CHARACTERIZATION OF THE NOVEL BROWN ADIPOCYTE CELL-LINE HIB 1B - ADRENERGIC PATHWAYS INVOLVED IN REGULATION OF UNCOUPLING PROTEIN GENE-EXPRESSION

The HIB 1B cell line, derived from a brown fat tumor of a transgenic m ouse, is the first established brown adipocyte cell line capable of ex pressing the brown fat-specific mitochondrial uncoupling protein (UCP) . UCP gene expression, which was virtually undetectable under basic co nditions, was stimulated by acute catecholamine or cyclic AMP treatmen t to levels comparable to primary cultures of brown adipocytes. Elevat ion of UCP mRNA levels following stimulation was very rapid but transi ent, decreasing after about 4 hours with a half-life between 9 and 13 hours. Immunoblotting showed the presence of UCP in HIB 1B mitochondri a, but expression was much lower than observed in BAT or primary cultu res of brown adipocytes. Upon transfection of HIB 1B cells with a repo rter gene containing the UCP promoter, the activity of the transgene w as regulatable by cAMP and norepinephrine. Investigation of the possib le adrenergic receptors involved in UCP stimulation showed that specif ic beta 3-adrenergic agonists were much less effective than nonspecifi c beta-adrenergic agonists and that mRNA levels of the atypical, fat-s pecific beta 3-adrenoceptor were lower than those observed in brown ad ipocytes differentiated in primary culture. From pharmacological evide nce we conclude that beta 3-adrenergic receptors account for approxima tely 30-40% of catecholamine induced UCP gene stimulation, whereas abo ut 60-70% is stimulated via the classical beta 1/2 adrenergic pathway. We conclude that HIB 1B cells represent a functional system for the s tudy of mechanisms related to brown adipose thermogenesis.