No abstract

This study compared the effects of centrally administered oxytocin (OT) and arginine vasopressin (AVP) on partner preference formation and social contact in male and female prairie voles (Microtus ochrogaster). After 1 hr of cohabitation and pretreatment with either AVP or OT, both males and females exhibited increased social contact and significant preference for the familiar partner. After pretreatment with either an OT receptor antagonist (OTA) or an AVP (Via) receptor antagonist (AVPA), neither OT nor AVP induced a partner preference. In addition, treatment with OT+OTA or AVP+AVPA was associated with low levels of social contact in both sexes. Either AVP or OT is sufficient to facilitate social contact if either the OT or AVP receptor is available. However, the formation of partner preferences may require access to both AVP and OT receptors.

Prairie voles (Microtus ochrogaster) are monogamous mammals that form male-female pair bonds. Partner preference formation, one component of the pair bond in prairie voles, occurs following male-female cohabitation and is facilitated by mating. The peptide hormone oxytocin is released during physical contact and particularly following vaginal stimulation. Oxytocin has been implicated in mother-infant bond formation. The present study tested the hypothesis that oxytocin participates in the partner preference component of pair bond formation in adult prairie voles. Ovariectomized female prairie voles were implanted with osmotic mini-pumps releasing oxytocin (1 -1 00 ng/h) or artificial cerebrospinal fluid (CSF). Pumps were implanted intracerebroventricularly or subcutaneously and females then were housed for 6 h with a male partner, followed by a preference test in which females could elect to spend time with either the partner or an unfamiliar male. Females in groups that received centrally-administered oxytocin (1 0 or 100 ng/h), but not CSF, exhibited a significant preference for the partner present during infusion. The induction of a partner preference after oxytocin administration appeared specific for central oxytocin pathways as peripheral oxytocin administration was ineffective. Moreover, central administration of a selective oxytocin receptor antagonist inhibited the behavioral effect of exogenous oxytocin. These results suggest that oxytocin may be one factor contributing to the development of partner preferences in this monogamous rodent.

The M2 protein of the influenza virus conducts protons into the virion under external acidic pH. The proton selectivity of the tetrameric channel is controlled by a single histidine (His(37)), whereas channel gating is accomplished by a single tryptophan (Trp(41)) in the transmembrane domain of the protein. Aromatic interaction between these two functional residues has been previously observed in Raman spectra, but atomic-resolution evidence for this interaction remains scarce. Here we use high-resolution solid-state NMR spectroscopy to determine the side-chain conformation and dynamics of Trp(41) in the M2 transmembrane peptide by measuring the Trp chemical shifts, His(37)-Trp(41) distances, and indole dynamics at high and low pH. The interatomic distances constrain the Trp41 side-chain conformation to trans for χ1 and 120-135° for χ2. This t90 rotamer points the Nε1-Cε2-Cζ2 side of the indole toward the aqueous pore. The precise χ1 and χ2 angles differ by ∼20° between high and low pH. These differences, together with the known changes in the helix tilt angle between high and low pH, push the imidazole and indole rings closer together at low pH. Moreover, the measured order parameters indicate that the indole rings undergo simultaneous χ1 and χ2 torsional fluctuations at acidic pH, but only restricted χ1 fluctuations at high pH. As a result, the Trp(41) side chain periodically experiences strong cation-π interactions with His(37) at low pH as the indole sweeps through its trajectory, whereas at high pH the indole ring is further away from the imidazole. These results provide the structural basis for understanding how the His(37)-water proton exchange rate measured by NMR is reduced to the small proton flux measured in biochemical experiments. The indole dynamics, together with the known motion of the imidazolium, indicate that this compact ion channel uses economical side-chain dynamics to regulate proton conduction and gating.

The influenza M2 protein not only forms a proton channel but also mediates membrane scission in a cholesterol-dependent manner to cause virus budding and release. The atomic interaction of cholesterol with M2, as with most eukaryotic membrane proteins, has long been elusive. We have now determined the cholesterol-binding site of the M2 protein in phospholipid bilayers using solid-state NMR spectroscopy. Chain-fluorinated cholesterol was used to measure cholesterol proximity to M2 while sterol-deuterated cholesterol was used to measure bound-cholesterol orientation in lipid bilayers. Carbon-fluorine distance measurements show that at a cholesterol concentration of 17 mol%, two cholesterol molecules bind each M2 tetramer. Cholesterol binds the C-terminal transmembrane (TM) residues, near an amphipathic helix, without requiring a cholesterol recognition sequence motif. Deuterium NMR spectra indicate that bound cholesterol is approximately parallel to the bilayer normal, with the rough face of the sterol rings apposed to methyl-rich TM residues. The distance- and orientation-restrained cholesterol-binding site structure shows that cholesterol is stabilized by hydrophobic interactions with the TM helix and polar and aromatic interactions with neighboring amphipathic helices. At the 1:2 binding stoichiometry, lipid P spectra show an isotropic peak indicative of high membrane curvature. This M2-cholesterol complex structure, together with previously observed M2 localization at phase boundaries, suggests that cholesterol mediates M2 clustering to the neck of the budding virus to cause the necessary curvature for membrane scission. The solid-state NMR approach developed here is generally applicable for elucidating the structural basis of cholesterol's effects on membrane protein function.