Waldherr et al. 10.1073/pnas.0705860104. |
Fig. 4. Anxiolytic effect of mating in males in the light phase. (a) Male Wistar rats were tested on the elevated plus maze 30 min after termination of a 30-min mating period performed in the light phase (i.e., 6-8 h after lights on). Adult male rats were either single-housed (yellow) or exposed to a nonreceptive (nonmated group; orange) or an estrogen/progesterone-primed (mated group; red) female rat in their home cage. Reduced anxiety levels in mated males were confirmed by increased exploration of the open arms of the plus maze. The locomotor activity as reflected by the number of entries into the closed arms was not significantly altered by mating in the light phase. Group size was as follows: single-housed, n = 12; nonmated, n = 26; mated, n = 10. Data represent means + SEM. *, P < 0.05 versus single-housed and nonmated males; #, P < 0.05 versus single-housed males after ANOVA. (b) The anxiolytic effect of mating did not correlate with the numbers of intromissions recorded during the 30-min mating period (mated animals from a and Fig. 1a).
Fig. 5. OT concentrations in individual microdialysates sampled from outside the hypothalamic PVN. (a) Two 15-min microdialysates were sampled during single-housing (dialysates 1 and 2), during the presence of a receptive female behind a perforated wall (dialysates 3 and 4), during mating with the female (dialysates 5 and 6), and after removal of the female from the male's home cage (dialysates 7 and 8). Histological verification revealed placement of microdialysis probes outside the PVN. The colors in a correspond to the respective color of microdialysis probe location in b. (b) Schematic drawings of the rat brain at the level of the hypothalamus and the PVN at two anterior-posterior planes (at bregma -0.8 and -1.5 mm) (3) where outsiders were located. The colored bars indicate the placement of microdialysis probes. 3V, third ventricle.
SI Text
Surgical Procedures.
Stereotaxic implantation of the microdialysis probe and the guide cannula for intracerebroventricular (icv) infusion were performed in male rats under isoflurane anesthesia as described in refs. 1 and 2. For monitoring OT release within the PVN, a U-shaped microdialysis probe (dialysis membrane: molecular cutoff of 18 kDa; Haemophan; Gambro Dialysatoren, Hechingen, Germany) was implanted into the right PVN (1.5 mm caudal to bregma, 1.8 mm lateral to midline, 9.1 mm beneath the surface of the skull, angle of 10° to avoid sagittal sinus damage) (3). For icv infusion of the receptor antagonists, a guide cannula (21 G, 12 mm long) was implanted with its tip resting 2 mm above the right lateral ventricle (1.0 mm caudal to bregma, 1.6 mm lateral to midline, 1.8 mm beneath the surface of the skull). The microdialysis probe and the icv guide cannula (closed with a stylet) were secured in place with dental cement to two stainless-steel screws inserted into the skull. After surgery, the rats received 30 ml of antibiotics s.c. (Baytril; Bayer Vital, Leverkusen, Germany). After surgery, animals were kept individually in transparent experimental cages (24 ´40 ´ 36 cm polycarbon walls) allowing detailed behavioral observation. They were handled daily to reduce nonspecific stress responses during the experiment. The correct placement of the microdialysis probe within the PVN and of the icv infusion was verified after termination of experiment on 20-mm cryocut slices by Nissl staining and by icv infusion of Evans Blue dye (5 ml), respectively.Intracerebral Microdialysis.
Microdialysates were sampled at 4.5 ml/min (sterile Ringer's solution, pH 7.4) into 1.5-ml Eppendorf tubes containing 10 ml of 0.1 M HCl and immediately stored at -80°C (1). Histological verification of placement of microdialysis probe was performed on Nissl-stained 40-mm cryocut sections.icv Infusion.
The receptor antagonist was slowly infused into the lateral ventricle of the well handled, freely moving animal in the home cage by using a 27-gauge icv cannula attached to a microliter syringe via 70-cm PE-50 tubing and inserted into the previously implanted guide cannula. After infusion, the icv cannula was kept in place for 30 s to allow substance diffusion. Both the oxytocin receptor and vasopressin V1a receptor antagonists were the generous gift of Dr. M. Manning (Toledo, OH).1. Neumann I, Russell JA, Landgraf R (1993) Neuroscience 53:65-75.
2. Neumann ID, Wigger A, Torner L, Holsboer F, Landgraf R (2000) J Neuroendocrinol 12:235-243.
3. Paxinos G, Watson C (1998) The Rat Brain in Stereotaxic Coordinates (Academic, Sydney).