Behavioral endocrinology of male rats with periodic sexual contacts with exclusive or varied females

Sexual contact keys a profound series of acute and chronic changes in males that, presumably, are orchestrated by acute pulsatile release of hormones. An experimental paradigm is reported in which male rats were paired periodically with either the same or different estrous females to receive identical amounts of copulatory experiences. Results confirmed the hypothesis that exposure to an unfamiliar female would induce a different endocrine response which would be reflected in various androgen-sensitive systems. The "successively polygynous" males showed more sexual behavior than "monogamous" males, and their respective females solicited the males differently, as well. Circulating levels of testosterone were higher immediately after sexual contact with an unfamiliar than with a familiar female partner. There were no differences in testosterone titers among the groups when the animals were killed at either 2 or 7 weeks after the final copulatory experiences. Yet, necropsies at 2 weeks postcopulation revealed that primary and secondary sex structures from polygynous males clearly were larger. Differences between the two experimental groups were reduced after 7 weeks of sexual rest, yet, polygnous males continued to show a different structural profile than the other groups. Conclusions were that males may experience greater activation of androgen-sensitive behavior and physiology following qualitatively different sexual contacts.     

Studies on some specific Ap4A-degrading enzymes with the use of various methylene analogues of P1P4-bis-(5'',5''''''-adenosyl) tetraphosphate.

Six new methylenephosphonate analogues of P1P4-bis-(5',5'-adenosyl) tetraphosphate, Ap4A, having P2-P3 carbon bridges CF2, CCl2 and CH2CH2 or P1-P2 and P3-P4 carbon bridges CF2, CCl2 and CH2CH2 in the tetraphosphate chain, were examined as substrates or inhibitors for two specific Ap4A-degrading enzymes: (asymmetrical) Ap4A hydrolase (EC 3.6.1.17) from yellow-lupin seeds and (symmetrical) Ap4A hydrolase (EC 3.6.1.41) from Escherichia coli. All analogues in which the central oxygen atom was replaced by a stable carbon bridge were hydrolysed by the asymmetrical hydrolase (CF2 greater than CCl2 greater than O greater than CHBr greater than CH2 greater than CH2CH2). As expected, these analogues were not hydrolysed by the symmetrical hydrolase, which was also unable to act on analogues having P1-P2 and P3-P4 carbon bridges.     

Evaluation of algorithms for ratio imaging in fluorescence microscopy

Ratio imaging in fluorescence microscopy is used in measuring parameters such as pH, pCa, cytoplasmic porosity, and the relative concentration of fluorescent analogs within single cells. The fastest method for ratio imaging is to use lookup tables on special-purpose image processors. Since lookup tables store integers in integer addresses, using a lookup table will generate rounding errors. The magnitude of the error will depend on the transformation performed and on the number of levels used in the lookup table. We examined ratio imaging by lookup table and computed the errors generated by both inversion and log subtraction methods. Both uniformly fluorescing fields and fluorescing cell images were employed to provide data for use in confirming our calculations and illustrating both the magnitude and spatial incidence of errors. It is shown that, through proper design of lookup tables, a significant reduction can be made in the errors generated in comparison with common methods available in most image processors.     

The unique insert of cellular and viral fms protein tyrosine kinase domains is dispensable for enzymatic and transforming activities.

The receptors for colony stimulating factor-1 (CSF-1), platelet derived growth factor and the c-kit protein tyrosine kinase (PTK) contain within their catalytic domains a stretch of 60-100 residues, largely unrelated in sequence, with no counterpart in other PTKs. Of the 64 amino acids within this kinase insert, 58 were deleted from the mouse CSF-1 receptor by oligonucleotide-directed mutagenesis. The mutant CSF-1 receptor was not markedly affected in its kinase activity, post-translational processing or its ability to induce autocrine transformation of NIH 3T3 mouse fibroblasts. Similarly, retention of kinase and transforming activities were observed following deletion of part or all of the kinase insert from the v-fms oncoprotein. The c- and v-fms kinase inserts were probed using monoclonal and polyclonal antibodies and were found to be highly antigenic. Two monoclonal antibodies raised to the v-fms cytoplasmic domain both recognized epitopes within the insert, and bound enzymatically active v-fms glycoproteins. These results indicate that the fms kinase insert is located on the surface of the protein and folds separately from the rest of the catalytic domain, but is not required for the biological activity of fms PTKs ectopically expressed in mouse fibroblasts. The insert may therefore play a specific function in cells such as monocytes and trophoblasts that normally express the CSF-1 receptor.     

Allogrooming, partner choice, and dominance in male anubis baboons

This study was designed to test the hypothesis that among unrelated male baboons (Papio cynocephalus anubis) in single-gender social groups there is no significant association between dominance status and allogrooming performance. The hypothesis was tested using behavioral measures obtained by focal animal sampling techniques. The results indicate that unrelated male baboons established well-defined linear dominance hierarchies, formed allogrooming relationships with one another, and exhibited a nonrandom distribution of allogrooming; however, there were no significant relationships between dominance rank and the frequency of allogrooming. We further tested our results by grouping individuals into three dominance status classes (high, middle, and low) and comparing the classes. Analysis of variance demonstrated no significant differences in rates of allogrooming by dominance class. These results suggest that dominance did not account for the variation in observed allogrooming behavior: Dominance status did not appear to determine the frequency with which animals groomed others, the number of grooming partners, or frequency of grooming that any individual received in comparison to that performed. High-ranking animals did not have significantly more grooming partners than low-ranking animals, and there appeared to be little competition within the groups for subordinates to groom high-ranking animals. When age, kinship, and group tenure are controlled, performance and reception of allogrooming are not strongly associated with dominance in single-gender social groups of male anubis baboons.     

Fluorescence energy transfer between cysteine 199 and cysteine 343: evidence for MgATP-dependent conformational change in the catalytic subunit of cAMP-dependent protein kinase

The catalytic subunit of cAMP-dependent protein kinase has two cysteine residues, Cys 199 and Cys 343, which are protected against alkylation by MgATP [Nelson, N. C., & Taylor, S. S. (1981) J. Biol. Chem. 256, 3743]. While Cys 199 is in close proximity to the active site of the catalytic subunit and is probably directly protected against alkylation by MgATP, the mechanism by which MgATP prevents alkylation of Cys 343 is unclear. To determine whether MgATP directly protects Cys 343 from alkylation by being in close proximity to both Cys 199 and the MgATP binding site, fluorescence resonance energy transfer techniques were used to measure the distance between Cys 199 and Cys 343. Two different donor-acceptor pairs containing 4-[N-[(iodoacetoxy)ethyl]-N-methylamino]-7-nitrobenz-2-oxa-1,3-diazole at Cys 199 as the acceptor and either 3,6,7-trimethyl-4-(bromomethyl)-1,5-diazabicyclo[3.3.0]octa-3,6-diene-2, 8- dione or N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl)ethylenediamine at Cys 343 as the donor were prepared following the method described in the preceding paper [First, E. A., & Taylor, S. S. (1989) Biochemistry (preceding paper in this issue)]. From the efficiencies of fluorescence resonance energy transfer for each donor-acceptor pair, the distance between Cys 199 and Cys 343 was estimated to be between 31 and 52 A. Since Cys 199 is close to the MgATP binding site and since MgATP cannot extend beyond a distance of 16 A, it is unlikely that Cys 343 at a distance of at least 31 A from Cys 199 is in direct contact with the bound nucleotide.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective modification of the catalytic subunit of cAMP-dependent protein kinase with sulfhydryl-specific fluorescent probes

The catalytic subunit of cAMP-dependent protein kinase contains only two cysteine residues, and the side chains of both Cys 199 and Cys 343 are accessible. Modification of the catalytic subunit by a variety of sulfhydryl-specific reagents leads to the loss of enzymatic activity. The differential reactivity of the two sulfhydryl groups at pH 6.5 has been utilized to selectively modify each cysteine with the following fluorescent probes: 3,6,7-trimethyl-4-(bromomethyl)-1,5-diazabicyclo[3.3.0]octa-3,6-diene- 2,8-dione, N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl)ethylenediamine, and 4-[N-[(iodoacetoxy)ethyl]-N-methyl-amino]-7-nitrobenz-2-oxa-1,3-diazole. The most reactive cysteine is Cys 199, and exclusive modification of this residue was achieved with each reagent at pH 6.5. Modification of Cys 343 required reversible blocking of Cys 199 with 5,5'-dithiobis(2-nitrobenzoic acid) followed by reaction of Cys 343 with the fluorescent probe at pH 8.3. Treatment of this modified catalytic subunit with reducing reagent restored catalytic activity by unblocking Cys 199. In contrast, catalytic subunit that was selectively labeled at Cys 199 by the fluorescent probes was catalytically inactive. Even though Cys 199 is presumably close to the interaction site between the regulatory subunit and the catalytic subunit, all of the modified C-subunits retained the capacity to aggregate with the type II regulatory subunit in the absence of cAMP, and the resulting holoenzymes were dissociated in the presence of cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)