Estradiol-17# administration to young (10-to 12-week-old) rabbits to produce the "estrogen-dominated" uterus increased the uterine contractile response to both oxjtocin and methacholine in vitro. In "progesterone-dominatel" uteri, obtained from rabbits that received progesterone for 4 days after estrogen pretreatment, the contractile response to oxytocin in vitro was selectively abolished; the response to methacholine was unaffected. Parallel changes were observed in the concentration (but not affinity) of specific sites in uterine microsomal membranes that bind 1111oxytocin with selectivity features expected for oxytocin receptors. fluence on the affinity of these sites. A sublethal dose of actinomycin D, administered over a 24-hr period to rabbits pretreated with estradiol for 4 days, likewise reduced specific oxytocin binding; additive effects were not observed when progesterone and actinomycin D were administered together. These results suggest that the regulatory effects of estrogens and progesterone upon the rabbit uterine contractile response to oxytocin are achieved, at least in part, by the opposing actions of these steroids in regulating the number of oxytocin receptors in smooth muscle cells. Estradiol increased the concentration of uterine oxytocin receptors; the maintenance of high receptor levels appears to depend upon the continuous de novo synthesis of oxytocin receptors. In contrast, progesterone, like actinomycin D, appears to act at the nuclear locus to repress synthesis of oxytocin receptors.

No abstract

Four highly purified synthetic peptides encompassing segments of the 68-86 region [for the numbering system used, see Eylar, E. H., Brostoff, S., Hashim, G., Caccam, J. & Burnett, P. (1971) J. Biol. Chem. 246, 5770-5784] of myelin basic protein (MBP), a region known to induce experimental allergic encephalomyelitis (EAE) in Lewis rats, were used to define and compare structure-function relationships between the primary structure of the 68-86 sequence and the three following biological activities: induction of EAE in Lewis rats, stimulation of T lymphocytes in vitro as measured by augmented cellular transfer of EAE to syngeneic recipients, and lymphocyte proliferation, as measured by [3Jthymidine incorporation. Guinea pig (GP) MBP was approximately 60 or 1500 times more active than the GP68-84 (Y G S L P Q K S Q R S Q D E N; single-letter amino acid abbreviations) or the modified bovine (MB) 68-84 (Y G S L P Q K A Q R P Q D E N) peptides for induction of EAE, respectively. Furthermore, lymphocytes primed with either GPMBP, GP68,84, or MB68-84 crossreacted in vitro with either GPMBP, GP68-84, or MB68-84 for activation of lymphocyte transfer activity. In contrast, lymphocytes primed with either GP68-84 or MB68-84 exhibited antigen-specific proliferation in vitro exclusively in response to either GP or MB sequences, respectively. Neither GP75-84 (S Q R S Q D E N) nor GP75-86 (S Q R S Q D E N P V) induced EAE, activated lymphocytes for EAE transfer, or stimulated lymphocyte proliferation under conditions and doses tested. We conclude that (i) structurally distinct determinants, reflecting existence of functionally independent classes of antigen receptors, specify encephalitogenic and proliferative responses of primed lymphocytes and (it) determinants for EAE induction, cellular transfer of EAE, and lymphocyte proliferation include amino acid residues in the 68-74 (Y G S L P Q K) sequence ofGPMBP.Experimental allergic encephalomyelitis (EAE) is a cellmediated autoimmune response directed against autologous central nervous system myelin. Disease manifestations include clinical neurologic signs and perivascular infiltration of inflammatory mononuclear cells within the central nervous system (1, 2). Sensitization with myelin basic protein (MBP), a major protein constituent of central nervous system compact myelin, emulsified in complete Freund's adjuvant, will produce the full clinical and histological picture of EAE in a wide variety of animal species. In Lewis rats, sensitization with MBP in complete Freund's adjuvant induces the generation of lymphocytes in peripheral lymphoid organs with the capacity to adoptively transfer EAE into syngeneic Lewis recipients (3, 4). When lymphocytes sensitized to MBP/ complete Freund's adjuvant are cultured in medium containing MBP for 3-4 days, they exhibit an '100-fold increase in EAE transfer activity, as evidenced by transfer of the paralytic signs of EAE with as few as 107 activated lymphocytes, and a vigorous antigen-specific proliferative response, as evidenced by [3...

but the exo/endo ratio is still only about 30. This low ratio stands in striking contrast to the one observed for solvolyses,8 which is ~105 at 25°, and it is concluded that neighboring group assistance is energetically less favored in the gas phase than in solution. Two reasons probably do account for this: firstly, neighboring group migration is usually associated with some loss of rotational entropy for the reactant on ring closure to the transition state.9 This entropy loss, which impedes reactivity, will inevitably be more pronounced in the gas phase than in solution. Secondly, since neighboring group assistance depends, among other factors, critically upon the charge development at the migrating center, the driving force for migration in the gas phase is reduced.

We prepared nine analogues (1-9) of MCPA-D-Phe-Phe-Ile-Asn-Cys-Pro-Arg-Gly-NH2, [MCPA1, D-Phe2, Phe3, Ile4, Arg8]oxytocin (MCPA = beta-mercapto-beta,beta-pentamethylenepropionic acid), a potent antagonist of the rat uterotonic action of oxytocin (OT). We replaced D-Phe with D-Trp and made [MCPA1,D-Trp2,Phe3,Ile4,Arg8]OT (1), which had OT pA2 of 7.51, somewhat higher than that of the D-Phe2 antagonist which has OT pA2 = 7.35 in our rat uterotonic assay. Both compounds are equipotent as antagonists of [Arg8]vasopressin in the rat antidiuretic assay, with pA2 = 8.1. Other substitutions gave [MCPA1,D-Trp2,4-Cl-Phe3,Ile4,Arg8]OT, (2), OT pA2 7.44; [MCPA1,D-Trp2,Phe3,Ile4,3,4-dehydro-Pro7,Arg8]OT (3), OT pA2 = 7.42; [MCPA1,D-Trp2,Phe3,Arg8]OT (4), OT pA2 = 7.58; [MCPA1,D-Trp2,Phe3,Arg8,Gly9-NHEt]OT (5), OT pA2 = 7.49; [MCPA1,D-Trp2,Ile4,Arg8]OT (6), OT pA2 = 7.46; [MCPA1,D-Trp2,Val4,Arg8]OT (7), OT pA2 = 7.58; [MCPA1,D-Trp2,Thr4,Arg8]OT (8), OT pA2 = 7.48; and finally, [MCPA1,D-Trp2,Arg8]OT (9), which was a more potent and more selective OT antagonist, with OT pA2 = 7.77 in the uterotonic assay and ADH pA2 less than 5.9 in the antidiuretic assay and hence is an important lead for the design of OT antagonists.