Association of the HPA axis response to upcoming competition and shooting outcomes in elite junior shooting players.

Successful shooting performance in competition is reliant on several factors such as shooting techniques and competition-associated psychological stresses. This study examined the hypothalamus-pituitary-adrenal (HPA) axis response to upcoming competition and its association with shooting outcomes in elite junior shooting players. The cortisol awakening response (CAR) and dehydroepiandrosterone (DHEA) secretion after awakening were measured for two consecutive days (the day before and on the day of competition for the selection of national shooting team members) in 19 junior men and 21 junior women players, and the shooting scores of the individual players were obtained. The total cortisol secretion during the CAR period (CARauc) increased, but total DHEA secretion during the post-awakening period (Daucawk) decreased on the day of competition, compared with one day before competition. The CARauc was higher in women than in men players, whereas Daucawk was higher in men than in women players across the two consecutive days. Cortisol and DHEA levels were comparable between low-scored (below the mean scores for air pistol or air rifle players) and high-scored players one day before competition. However, the CARauc on the day of competition was higher and the variations in the CARauc and molar CARauc/Daucawk ratios across the two consecutive days were greater in low-scored than in high-scored men and women players. These results indicated that upcoming competition involves alterations of the CAR and DHEA secretion after the awakening period, and greater HPA response to the upcoming competition was adversely associated with shooting scores in junior shooting athletes.Lay summaryAn important upcoming competition was perceived as a strong stressor on awakening that induced alteration in CAR and DHEA secretion after the awakening period in elite shooting players. This study observed that, irrespective of their age and period of shooting practice, the HPA axis function on the day of competition was associated with shooting outcomes in elite shooting players.

Altered Hypothalamus-Pituitary-Adrenal Axis Function: A Potential Underlying Biological Pathway for Multiple Concurrent Symptoms in Patients With Advanced Lung Cancer.

OBJECTIVE: Patients with advanced cancer commonly experience multiple symptoms that present as groups or clusters. The present study aimed to examine whether hypothalamus-pituitary-adrenal (HPA) axis dysfunction underlies the concurrent multiple symptoms in patients with advanced cancer. METHODS: Patients' cortisol levels were determined in saliva samples collected after awakening (0, 30, and 60 minutes after awakening) and at nighttime (21:00-22:00 PM) from 46 patients with lung cancer (15.2% women), with a mean (standard deviation) age of 64.3 (9.2) years and 47 healthy participants (53.2% women; age = 62.0 [4.6] years). Cancer-related symptoms were measured using the M.D. Anderson Symptom Inventory (MDASI). RESULTS: Compared with healthy participants, patients showed a significantly reduced cortisol awakening response (F(1,364) = 46.2, p < .001) and had flatter diurnal slope of cortisol (larger ß values) (mean [standard error of the mean] = -0.64 [0.06] versus -0.18 [0.05], p < .001). Altered HPA axis function was significantly and adversely associated with performance status and burden of symptoms (all p values < .01). However, each MDASI item varied widely in the degree of association with the HPA axis function. Hierarchical clustering analysis based on Spearman's rank correlation with complete linkage identified that nausea was clustered with vomiting, numbness, and dry mouth, whereas the other nine MDASI core symptoms associated with altered HPA axis function were clustered together. CONCLUSIONS: Altered HPA axis function may be a possible biological pathway that can explain the concurrence of core symptoms in patients with advanced lung cancer.

Insulin directly regulates steroidogenesis via induction of the orphan nuclear receptor DAX-1 in testicular Leydig cells.

Testosterone level is low in insulin-resistant type 2 diabetes. Whether this is due to negative effects of high level of insulin on the testes caused by insulin resistance has not been studied in detail. In this study, we found that insulin directly binds to insulin receptors in Leydig cell membranes and activates phospho-insulin receptor-ß (phospho-IR-ß), phospho-IRS1, and phospho-AKT, leading to up-regulation of DAX-1 (dosage-sensitive sex reversal, adrenal hypoplasia critical region, on chromosome X, gene 1) gene expression in the MA-10 mouse Leydig cell line. Insulin also inhibits cAMP-induced and liver receptor homolog-1 (LRH-1)-induced steroidogenic enzyme gene expression and steroidogenesis. In contrast, knockdown of DAX-1 reversed insulin-mediated inhibition of steroidogenesis. Whether insulin directly represses steroidogenesis through regulation of steroidogenic enzyme gene expression was assessed in insulin-injected mouse models and high fat diet-induced obesity. In insulin-injected mouse models, insulin receptor signal pathway was activated and subsequently inhibited steroidogenesis via induction of DAX-1 without significant change of luteinizing hormone or FSH levels. Likewise, the levels of steroidogenic enzyme gene expression and steroidogenesis were low, but interestingly, the level of DAX-1 was high in the testes of high fat diet-fed mice. These results represent a novel regulatory mechanism of steroidogenesis in Leydig cells. Insulin-mediated induction of DAX-1 in Leydig cells of testis may be a key regulatory step of serum sex hormone level in insulin-resistant states.

Phosphoenolpyruvate carboxykinase and glucose-6-phosphatase are required for steroidogenesis in testicular Leydig cells.

Cyclic AMP (cAMP) induces steroidogenic enzyme gene expression and stimulates testosterone production in Leydig cells. Phosphoenolpyruvate carboxykinase (PEPCK) is expressed in Leydig cells, but its role has not been defined. In this study, we found that PEPCK and glucose-6-phosphatase (Glc-6-Pase) are increased significantly following cAMP treatment of mouse Leydig cells. Moreover, cAMP treatment increased recruitment of the cAMP-response element-binding transcription factor and decreased recruitment of the corepressor DAX-1 on the pepck promoter. Furthermore, cAMP induced an increase in ATP that correlated with a decrease in phospho-AMP-activated protein kinase (AMPK). In contrast, knockdown or inhibition of PEPCK decreased ATP and increased phospho-AMPK. Treatment with an AMPK activator or overexpression of the constitutively active form of AMPK inhibited cAMP-induced steroidogenic enzyme promoter activities and gene expression. Liver receptor homolog-1 (LRH-1) was involved in cAMP-induced steroidogenic enzyme gene expression but was inhibited by AMPK activation in Leydig cells. Additionally, inhibition or knockdown of PEPCK and Glc-6-Pase decreased cAMP-mediated induction of steroidogenic enzyme gene expression and steroidogenesis. Finally, pubertal mouse (8-week-old) testes and human chorionic gonadotropin-induced prepubertal mouse testes showed increased PEPCK and Glc-6-Pase gene expression. Taken together, these results suggest that induction of PEPCK and Glc-6-Pase by cAMP plays an important role in Leydig cell steroidogenesis.

Molecular cloning and expression of steroidogenic acute regulatory protein from bullfrog (Rana catesbeiana).

Steroidogenic acute regulatory protein (StAR) transfers cholesterol from the outer mitochondrial membrane to the inner membrane where the cytochrome P450 side chain cleavage enzyme (P450scc) resides. This process is the rate-limiting step in steroidogenesis. StAR cDNAs have been cloned and characterized from a range of different species. To investigate the role of StAR in the amphibian system, we cloned a full-length StAR cDNA from bullfrog (Rana catesbeiana) using reverse transcription polymerase chain reaction (RT-PCR) in conjunction with rapid amplification of cDNA ends (RACE). The putative full-length bullfrog StAR (bfStAR) cDNA was 1862 base pairs (bp) in length, and the longest open reading frame (ORF) encoded a protein of 284 amino acids. Amino acid sequence comparison showed that amphibian StAR has a high degree of sequence identity, ranging from 62% to 98%, with StAR proteins of other species. Similar to other species, bfStAR contained two conserved domains, the mitochondrial targeting domain and cholesterol-binding domain, in the N-terminus and C-terminus of the protein, respectively. Northern blot analysis and RT-PCR indicated that StAR mRNA is expressed in the gonads and adrenal gland. Transfection of green monkey kidney (COS-1) cells with an expression construct for bfStAR revealed that it encoded 34 and 27kDa proteins that were recognized by antiserum raised against the human StAR-related lipid transfer (START) domain.

Reduced testicular steroidogenesis in tumor necrosis factor-alpha knockout mice.

We previously demonstrated that the expression of Mullerian inhibiting substance (MIS) in Sertoli cells is downregulated by tumor necrosis factor alpha (TNF-alpha), which is secreted by meiotic germ cells, in mouse testes. Several studies have reported that MIS that is secreted by Sertoli cells inhibits steroidogenesis and, thus, the synthesis of testosterone in testicular Leydig cells. Here, we demonstrate that in TNF-alpha knockout testes, which show high levels of MIS, steroidogenesis is decreased compared to that in wild-type testes. The levels of testosterone and the mRNA levels of steroidogenesis-related genes were significantly lower after puberty in TNF-alpha knockout testes than in wild-type testes. Furthermore, the number of sperm was reduced in TNF-alpha knockout mice. Histological analysis revealed that spermatogenesis is also delayed in TNF-alpha knockout testes. In conclusion, TNF-alpha knockout mice show reduced testicular steroidogenesis, which is likely due to the high level of testicular MIS compared to that seen in wild-type mice.

Salivary cortisol and DHEA levels in the Korean population: age-related differences, diurnal rhythm, and correlations with serum levels.

PURPOSE: The primary objective of this study was to examine the changes of basal cortisol and DHEA levels present in saliva and serum with age, and to determine the correlation coefficients of steroid concentrations between saliva and serum. The secondary objective was to obtain a standard diurnal rhythm of salivary cortisol and DHEA in the Korean population. MATERIALS AND METHODS: For the first objective, saliva and blood samples were collected between 10 and 11 AM from 359 volunteers ranging from 21 to 69 years old (167 men and 192 women). For the second objective, four saliva samples (post-awakening, 11 AM, 4 PM, and bedtime) were collected throughout a day from 78 volunteers (42 women and 36 men) ranging from 20 to 40 years old. Cortisol and DHEA levels were measured using a radioimmunoassay (RIA). RESULTS: The morning cortisol and DHEA levels, and the age- related steroid decline patterns were similar in both genders. Serum cortisol levels significantly decreased around forty years of age (p < 0.001, when compared with people in their 20s), and linear regression analysis with age showed a significant declining pattern (slope=-2.29, t=-4.297, p < 0.001). However, salivary cortisol levels did not change significantly with age, but showed a tendency towards decline (slope=-0.0078, t=-0.389, p=0.697). The relative cortisol ratio of serum to saliva was 3.4-4.5% and the ratio increased with age (slope=0.051, t=3.61, p < 0.001). DHEA levels also declined with age in saliva (slope=-0.007, t=-3.76, p < 0.001) and serum (slope=-0.197 t=-4.88, p < 0.001). In particular, DHEA levels in saliva and serum did not start to significantly decrease until ages in the 40s, but then decreased significantly further at ages in the 50s (p < 0.001, when compared with the 40s age group) and 60s (p < 0.001, when compared with the 50 age group). The relative DHEA ratio of serum to saliva was similar throughout the ages examined (slop=0.0016, t=0.344, p=0.73). On the other hand, cortisol and DHEA levels in saliva reflected well those in serum (r=0.59 and 0.86, respectively, p < 0.001). The highest salivary cortisol levels appeared just after awakening (about two fold higher than the 11 AM level), decreased throughout the day, and reached the lowest levels at bedtime (p < 0.001, when compared with PM cortisol levels). The highest salivary DHEA levels also appeared after awakening (about 1.5 fold higher than the 11 AM level) and decreased by 11 AM (p < 0.001). DHEA levels did not decrease further until bedtime (p=0.11, when compared with PM DHEA levels). CONCLUSION: This study showed that cortisol and DHEA levels change with age and that the negative slope of DHEA was steeper than that of cortisol in saliva and serum. As the cortisol and DHEA levels in saliva reflected those in serum, the measurement of steroid levels in saliva provide a useful and practical tool to evaluate adrenal functions, which are essential for clinical diagnosis.

Effects of butyltin compounds on follicular steroidogenesis in the bullfrog(Rana catesbeiana).

The effects of butyltin compounds on follicular steroidogenesis in amphibians were examined using ovarian follicles of Rana catesbeiana. Isolated follicles were cultured for 18h in the presence and absence of frog pituitary homogenate (FPH) or various steroid precursors, and the steroid levels in the follicles or culture media were measured by radioimmunoassay (RIA). Among the butyltin compounds, tributyltin (TBT) strongly inhibited the FPH-induced synthesis of pregnenolone (P(5)), progesterone (P(4)) and testosterone (T). It also inhibited the conversion of P(5)-P(4) and T to estradiol-17ß(E(2)) and it partially suppressed the conversion of androstenedione (AD) to T, but not P(4) to 17α-hydroxyprogesterone (17α-OHP(4)). A high concentration of dibutyltin (DBT) also inhibited steroidogenesis by the follicles while monobutyltin (MBT) and tetrabutylin (TeBT) had no effect. These results suggest that the initial step of steroidogenesis (P(5) synthesis) and enzymes such as 3ß-HSD, 17ß-HSD and aromatase are inhibited by TBT or DBT. However, 17α-hydroxylase was not suppressed by TBT or the other butyltin compounds.

Effect of ascorbic acid supplementation on testicular steroidogenesis and germ cell death in cadmium-treated male rats.

Cadmium (Cd) is one of the environmental pollutants affecting various tissues and organs including testis. Harmful effect of Cd in testis is known to be germ cell degeneration and impairment of testicular steroidogenesis. Animals treated with high doses of Cd (0.2 and 0.3 mg/100g BW) showed a significant decrease in serum testosterone (T) level, but a significant induction of testicular lipid peroxidation levels. TUNEL assay showed that low doses of Cd (0.13 and 0.15 mg/100g BW) exhibited typical characteristics of apoptosis while high doses of Cd caused more necrosis than apoptosis. In contrast, supplementation with ascorbic acid reduced testicular lipid peroxidation levels. Ascorbic acid supplementation restored testicular 3beta-hydroxysteroiddehydrogenase (HSD) and 17beta-HSD enzyme activities, 3beta-HSD and cytochrome P450 side chain cleavage (P450(scc)) mRNA levels and serum T concentration to normal in Cd-administered rats. Moreover, administration of ascorbic acid prevented germ cell apoptosis as demonstrated by the reduced number of TUNEL-positive cells in germinal epithelium and inhibited Cd-induced necrosis. These results indicate that ascorbic acid have protective roles in vivo on the Cd-induced overall testicular damage including impaired steroidogenesis and germ cell death possibly through scavenging the reactive oxygen species generated by Cd administration.

Differential G protein coupling preference of mammalian and nonmammalian gonadotropin-releasing hormone receptors.

Recently, we have identified three distinct types of gonadotropin-releasing hormone receptor (GnRHR) in the bullfrog (designated bfGnRHR-1, bfGnRHR-2, and bfGnRHR-3). In the present study, we compared G protein coupling preference of mammalian and nonmammalian GnRHRs. In a transient expression system, stimulation of either bfGnRHRs or rat GnRHR by GnRH significantly increased both inositol phosphates (IP) and cAMP productions, but ratios of IP to cAMP induction levels were quite different among the receptors, indicating differential G protein coupling preference. Using cAMP-dependent protein kinase A (PKA)-specific (CRE-luc) or protein kinase C (PKC)-specific reporter (c-fos-luc) systems, we further examined G(s) and G(q/11) coupling preference of these GnRHRs. Since activities of CRE-luc and c-fos-luc were highly dependent on cell types, GnRH-induced CRE-luc or c-fos-luc activity was normalized by forskolin-induced CRE-luc or 12-O-tetradecanoylphenol-13-acetate (TPA)-induced c-fos-luc activity, respectively. This normalized result indicated that bfGnRHR-2 couples to G(s) more actively than G(q/11), while bfGnRHR-1 and -3 couple to G(s) and G(q/11) with similar strength. However, the rat GnRHR appeared to couple to G(q/11) more efficiently than G(s). This study was further confirmed by an experiment in which GnRH augmented CRE-driven luciferase activity in alphaT3-1 cells when CRE-luc was cotransfected with bfGnRHRs but not with vehicle or rat GnRHR. Collectively, these results indicate that mammalian and nonmammalian GnRHRs may induce diverse cellular and physiological responses through differential activation of PKA and PKC signaling pathways.

Antiandrogenic effects of bisphenol A and nonylphenol on the function of androgen receptor.

By using a yeast detection system for androgenic and antiandrogenic effects of chemicals, we identified bisphenol A (BPA) and nonylphenol (NP) as antiandrogens. In this study, we report molecular mechanisms for the antiandrogenic action of BPA and NP. In the ARhLBD-activating signal cointegrator 1 (ASC1) yeast two-hybrid system, which reflects the androgen-dependent interaction between androgen receptor (AR) and its coactivator, ASC1, BPA and NP acted as potent AR antagonists comparable to a known strong antagonist, cyproterone acetate. Ligand competition assays revealed that [3H]5alpha-dihydroxytestosterone (DHT) binding to AR is inhibited a maximum of 30 and 40% at approximately 5 nM of NP and 50 nM of BPA, respectively. In addition, the nuclear translocation of green fluorescent protein (GFP)-AR fusion protein in the presence of testosterone was affected by the addition of BPA and NP, which cause rather dispersed distribution of GFP-AR between the nuclear and the cytoplasmic compartments. Furthermore, in transient transfection assays, BPA and NP inhibited androgen-induced AR transcriptional activity. Taken together, the results suggest that BPA and NP affect multiple steps of the activation and function of AR, thereby inhibiting the binding of native androgens to AR, AR nuclear localization, AR interaction with its coregulator, and its subsequent transactivation. These data may help us better understand the biological alterations induced by these environmental compounds.

Transforming growth factor-ß1 signaling represses testicular steroidogenesis through cross-talk with orphan nuclear receptor Nur77.

Transforming growth factor- ß1 (TGF-ß1) has been reported to inhibit luteinizing hormone (LH) mediated-steroidogenesis in testicular Leydig cells. However, the mechanism by which TGF-ß1 controls the steroidogenesis in Leydig cells is not well understood. Here, we investigated the possibility that TGF-ß1 represses steroidogenesis through cross-talk with the orphan nuclear receptor Nur77. Nur77, which is induced by LH/cAMP signaling, is one of major transcription factors that regulate the expression of steroidogenic genes in Leydig cells. TGF-ß1 signaling inhibited cAMP-induced testosterone production and the expression of steroidogenic genes such as P450c17, StAR and 3ß-HSD in mouse Leydig cells. Further, TGF-ß1/ALK5 signaling repressed cAMP-induced and Nur77-activated promoter activity of steroidogenic genes. In addition, TGF-ß1/ALK5-activated Smad3 repressed Nur77 transactivation of steroidogenic gene promoters by interfering with Nur77 binding to DNA. In primary Leydig cells isolated from Tgfbr2flox/flox Cyp17iCre mice, TGF-ß1-mediated repression of cAMP-induced steroidogenic gene expression was significantly less than that in primary Leydig cells from Tgfbr2flox/flox mice. Taken together, these results suggest that TGF-ß1/ALK5/Smad3 signaling represses the expression of steroidogenic genes via the suppression of Nur77 transactivation in testicular Leydig cells. These findings may provide a molecular mechanism involved in the TGF-ß1-mediated repression of testicular steroidogenesis.

ERα/E2 signaling suppresses the expression of steroidogenic enzyme genes via cross-talk with orphan nuclear receptor Nur77 in the testes.

Estrogen receptor alpha (ERα) has been reported to affect steroidogenesis in testicular Leydig cells, but its molecular mechanism remains unclear. Here, we investigate the effect of estrogen and ERα on Nur77, a major transcription factor that regulates the expression of steroidogenic enzyme genes. In MA-10 Leydig cells, estradiol (E2) treatment, and interestingly ERα overexpression, suppressed the cAMP-induced and Nur77-activated promoter activity of steroidogenic enzyme genes via the suppression of Nur77 transactivation. ERα physically interacted with Nur77 and inhibited its DNA binding activity. In addition, ERα/E2 signaling decreased Nur77 protein levels. Consistent with the above results, the testicular testosterone level was higher in Leydig cell-specific ERα knock-out mice (ERα(flox/flox)Cyp17iCre) than in wild-type mice (ERα(flox/flox)). Taken together, these results suggest that ERα/E2 signaling controls the Nur77-mediated expression of steroidogenic enzyme genes in Leydig cells. These findings may provide a mechanistic explanation for the local regulation of testicular steroidogenesis by estrogenic compounds and ERα.

Cortisol awakening response and nighttime salivary cortisol levels in healthy working Korean subjects.

PURPOSE: Cortisol awakening response (CAR) and nighttime cortisol levels have been used as indices of adrenocortical activity. However, population-based statistical information regarding these indices has not been provided in healthy subjects. This study was carried out to provide basic statistical information regarding these indices. MATERIALS AND METHODS: Cortisol levels were measured in saliva samples collected immediately upon awakening (0 min), 30 min after awakening and in the nighttime on two consecutive days in 133 healthy subjects. RESULTS: We determined the mean [standard deviation (SD)], median (interquartile range) and 5th-95th percentile range for each measure and auxiliary indices for CAR, i.e., the secreted cortisol concentration within 30 min of awakening (CARscc) and absolute and relative increases in cortisol level within 30 min of awakening (CARi and CARi%, respectively). We also determined these values for auxiliary indices derived from nighttime cortisol level, i.e., the ratio of cortisol level 30 min after awakening (CA(30 min)) to nighttime level (CA(30 min)/NC), as well as absolute and relative decreases in cortisol levels from CA(30 min) to nighttime (DCd and DCd%, respectively). We found no significant differences in cortisol level for any time point or in auxiliary indices between collection days, genders and ages. CONCLUSION: The provided descriptive information and statistics on the CAR and nighttime cortisol level will be helpful to medical specialists and researchers involved in hypothalamus- pituitary-adrenal axis assessment.

Day-to-day differences in cortisol levels and molar cortisol-to-DHEA ratios among working individuals.

PURPOSE: The present study was carried out to determine day-to-day differences in cortisol levels and the molar cortisol-to-dehydroepiandrosterone (DHEA) ratio (molar C/D ratio) in working subjects. MATERIALS AND METHODS: The cortisol and DHEA levels were measured from saliva samples collected 30 minutes after awakening for 7 consecutive days in full-time working subjects that worked Monday through Saturday. To determine the day-to-day differences within subjects, the collected data was analyzed using variance (ANOVA) for a randomized complete block design (RCBD). RESULTS: The cortisol levels from samples collected 30 minutes after awakening on workdays were similar to each other, but were significantly different from the cortisol levels on Sunday. The DHEA levels were not significantly different between the days of week. The DHEA levels on Monday and Tuesday were relatively lower than the levels on the other weekdays. The DHEA levels on Thursday and Friday were relatively higher than the other days. The molar C/D ratios on Sunday were significantly lower than those on workdays. The molar C/D ratios on Monday and Tuesday were significantly higher than those on Wednesday or other workdays. CONCLUSION: The cortisol levels and the molar C/D ratios demonstrate differences in adrenocortical activities between workdays and non-workdays, but the molar C/D ratio additionally represents differences in adrenocortical status between the first two workdays and other workdays. Thus, it is possible that the day-to-day differences in the cortisol levels and the molar C/D ratio represent the adrenal response to upcoming work-related stress.

Identification of amino acid residues that direct differential ligand selectivity of mammalian and nonmammalian V1a type receptors for arginine vasopressin and vasotocin. Insights into molecular coevolution of V1a type receptors and their ligands.

Arginine vasotocin (VT) is the ortholog in all nonmammalian vertebrates of arginine vasopressin (AVP) in mammals. We have previously cloned an amphibian V1atype vasotocin receptor (VT1R) that exhibited higher sensitivity for VT than AVP, while the mammalian V1a type receptor (V1aR) responded better to AVP than VT. In the present study, we identified the amino acid residues that confer differential ligand selectivity for AVP and VT between rat V1aR and bullfrog VT1R (bfVT1R). A chimeric rat V1aR having transmembrane domain (TMD) VI to the carboxyl-terminal tail (C-tail) of bfVT1R showed a reverse ligand preference for AVP and VT, whereas a chimeric VT1R with TMD VI to the C-tail of rat V1aR showed a great increase in sensitivity for AVP. A single mutation (Ile(315(6.53)) to Thr) in TMD VI of V1aR increased the sensitivity for VT, while a single mutation (Phe(313(6.51)) to Tyr or Pro(334(7.33)) to Thr) reduced sensitivity toward AVP. Interestingly the triple mutation (Phe(313(6.51)) to Tyr, Ile(6.53) to Thr, and Pro(7.33) to Thr) of V1aR increased sensitivity to VT but greatly reduced sensitivity to AVP, behaving like bfVT1R. Further, like V1aR, a double mutant (Tyr(306(6.51)) to Phe and Thr(327(7.33)) to Pro) of bfVT1R showed an increased sensitivity to AVP. These results suggest that Phe/Tyr(6.51), Ile/Thr(6.53), and Pro/Thr(7.33) are responsible for the differential ligand selectivity between rat V1aR and bfVT1R. This information regarding the molecular interaction of VT/AVP with their receptors may have important implications for the development of novel AVP analogs.