Endogenous oxytocin levels in extracted saliva elevates during breastfeeding correlated with lower postpartum anxiety in primiparous mothers

doi:10.21203/rs.3.rs-1421477/v1

Download PDF

Research Article

Endogenous oxytocin levels in extracted saliva elevates during breastfeeding correlated with lower postpartum anxiety in primiparous mothers

https://doi.org/10.21203/rs.3.rs-1421477/v1

This work is licensed under a CC BY 4.0 License

You are reading this latest preprint version

Background

Breastfeeding in the early postpartum period is expected to have mental benefits for mothers; however, the underlying psychobiological mechanisms remain unclear. Previously, we hypothesized that the release of oxytocin in response to the suckling stimuli during breastfeeding would mediate a calming effect on primiparous mothers, and we examined salivary oxytocin measurements in primiparous mothers at postpartum day 4 using saliva samples without extraction, which was erroneous. Thus, further confirmation of this hypothesis with a precise methodology was needed.

Methods

We collected saliva samples at three time points to measure oxytocin in 24 primiparous mothers on postpartum day 2 (PD2) and 4 (PD4) across the breastfeeding cycle (before, during, and after breastfeeding). Salivary oxytocin levels using both extracted and unextracted method were measured and compared to show the qualitative differences. State and trait anxiety and clinical demographics were evaluated to determine their association with oxytocin changes.

Results

Breastfeeding elevated salivary oxytocin levels at PD2; however, there was no significant increase at PD4 in the extracted method. We found a weak but significant positive correlation between extracted and unextracted oxytocin levels during breastfeeding; there were no other significant positive correlations. Therefore, we used the extracted measurement index for subsequent analysis. We showed that the greater the increase in oxytocin during breastfeeding, the lower the state anxiety, but not trait anxiety. Mothers who exclusively breastfed at the 1-month follow-up had a slightly higher oxytocin change at PD2 compared to those who did not.

Conclusions

Breastfeeding in early postpartum days could be accompanied by frequent release of oxytocin and lower state anxiety, potentially contributing to exclusive breastfeeding.

calming effect

extraction

maternity blues

postpartum depression

Breastfeeding in the early postpartum period not only promotes the recovery of the reproductive organs of the mother but also enhances the immunity of the newborn and prevents infections and allergies [1–3]. Since the physical benefits to both the mother and newborn in the early postpartum period are considerable, breastfeeding during this period has been especially promoted worldwide through the “Ten steps to successful breastfeeding” proposed by the World Health Organization (WHO) [4]. In addition, early postpartum breastfeeding has a decisive influence on the persistence of breastfeeding following the newborn period. Breastfeeding also supports the interaction and bonding between mothers and infants. A longer breastfeeding duration was associated with greater maternal-sensitive responsiveness, more attachment security, and less attachment disorganization in the child [5]. There is also a large body of evidence of the psycho-physical benefits of persistent exclusive breastfeeding, such as better attachment formation between the mother and child, which may forestall child abuse and neglect, and the benefits to the child’s intelligence and development [6, 7]. However, few studies have examined its psychobiological basis whereby early postpartum breastfeeding provides mental benefits to mothers.

The neuroendocrine basis of breastfeeding is driven by prolactin and oxytocin (OT), which are secreted by the pituitary gland. Prolactin is involved in milk production, and OT stimulates milk ejection by contracting the myoepithelium surrounding the breast. In recent years, OT has attracted more attention as a neurotransmitter than for its classical properties as a hormone [8]. Intranasal OT has been shown to have psychoactive effects and increase trust, empathy, and other prosocial qualities, by directly affecting the brain [9]. Therefore, we hypothesized that early postpartum breastfeeding may be linked to a phenomenon that provides psychobiological benefits to mothers by specifically enhancing endogenous OT secretion and regulating the neural substrates that regulate cognition and emotion.

It has been well recognized that mothers’ plasma OT levels rise in response to the suckling stimulation during breastfeeding [10]. A study of plasma OT levels in breastfeeding mothers from 2 days to 11 weeks postpartum reported increased OT levels during breastfeeding [11, 12]. Additionally, plasma OT concentration increases within 1 min after the initiation of breastfeeding and returns to basal levels within 6 min after the end of breastfeeding, indicating that this increase in OT concentration is caused by breastfeeding [13]. However, no study has yet examined whether this endogenous OT increase by breastfeeding has a beneficial effect on the mother’s psychological state. While OT has been measured mostly using plasma, recent years have seen an expansion in the use of saliva, as salivary OT has been shown to correlate with plasma and cerebrospinal fluid OT [14–17]. However, when similar experiments were performed from the measurement of salivary OT, this phenomenon of increased OT concentration after breastfeeding did not always seem to be captured. For example, a study of 6-month postpartum mothers reported that salivary OT concentration during breastfeeding was highest 30 min before breastfeeding, decreased during breastfeeding, and increased slightly 30 min after the end of breastfeeding [18, 19]. Other groups have also conducted similar experiments and reported that they did not capture an increase in OT concentration [20]. In contrast, we conducted a similar experiment in first-time mothers on postpartum days 4–5, showing that salivary OT concentration increased during breastfeeding and was still higher than baseline at 30 min after breastfeeding [21]. We concluded that this might be because our experiment was conducted in the early postpartum period, unlike the other groups, and the neuroendocrine basis for the oxytocinergic system in mothers may be different. In addition, salivary OT that increased with breastfeeding was negatively correlated with maternal anxiety, indicating that endogenous OT secretion during early postpartum breastfeeding may have an anxiolytic effect [21]. However, our previous study was not able to eliminate the effect of false positives and had issues with precision of the measurement because saliva OT was measured using the unextracted method. In addition, anxiety was assessed using the anxiety subscale of the shortened version of the Profile of Mood States (POMS) [22], since we did not have a specific hypothesis for the anxiolytic effect at the time. The anxiety subscale of the POMS, which presumably assesses state anxiety, is also indistinguishable from trait anxiety. On the other hand, since the phase of salivary OT secretion can change at different periods after childbirth, as observed in other studies [18–20], the phase of OT secretion may differ during the first few days after childbirth. Furthermore, the symptoms of maternity blues are caused by a sudden decline in gonadal hormones after delivery, with a time lag of several days [23]. Given that the symptoms of maternity blues are transient, but in some cases, worsen the symptoms of postpartum depression [23], it is valuable to compare the effects of OT on the first few days postpartum.

In the present study, to test our hypothesis, we used a more sophisticated extraction method to measure salivary OT and assessed state and trait anxiety using a standard questionnaire State-Trait Anxiety Inventory (STAI) at two time points, postpartum days 2 and 4.

Participants

Twenty-four Japanese mothers hospitalized in the Miyamura Obstetrics and Gynecology Clinic, Nagasaki, participated in this study on postpartum days 2 (PD2) and 4 (PD4). They were recruited during pregnancy and were eligible if they met the following criteria [21]: (i) primiparous; (ii) currently exclusively breastfeeding their infants; and (iii) had no history of mental disorders, including depression and anxiety, and no current medication use or history of psychotropic medication use. We used the following exclusion criteria: (i) recent obstetrical complications involving the mothers or their infants, (ii) delivery of a preterm infant, or (iii) delivery by cesarean section. Among the 24 mothers, two were excluded because they provided insufficient saliva. Thus, 22 mothers were included in the final analysis. The clinical demographics of the participants are presented in Table 1. These participants were independent of the participants in our previous study [21].

Table 1

Clinical demographics of participants
	Primiparous mothers (n = 22)
Age (years)	30.6 ± 5.0 (19–40)
Gestational age (week)	39.7 ± 1.1 (36.0–41.4)
Duration of labor (min)	1025.0 ± 554.6 (378–2341)
Number of nipple pores PD2 PD4	2.4 ± 0.8 (1–4) 5.2 ± 2.3 (1–9)
Number of breastfeeding cycles per day PD1 PD2 PD3	2.6 ± 1.1 (0–4) 3.7 ± 1.0 (2–6) 4.8 ± 1.1 (3–7)
Nocturnal sleep time (h) PD1 to 2 PD3 to 4	6.8 ± 2.1 (2.5–12) 5.5 ± 1.4 (2–8)
Breast engorgement (severe / moderate / none) PD2 PD4	1 (4%) / 3 (14%) / 18 (82%) 5 (23%) / 15 (68%) / 2 (9%)
Induction medicine during labor (Yes / No)	6 (27%) / 16 (73%)
Uterine contraction medicine after labor (Yes / No)	2 (9%) / 20 (91%)
Perineotomy during labor (Yes / No)	2 (9%) / 20 (91%)
Duration of breastfeeding at the experiment (min) PD2 PD4	11.1 ± 3.5 (5–20) 17.3 ± 3.6 (7–20)
State anxiety (STAI-State) PD2 PD4	39.9 ± 7.7 (27–54) 36.4 ± 9.1 (22–59)
Trait anxiety (STAI-Trait)	41.1 ± 7.3 (27–54)
Breastfeeding style at 1-month follow-up (exclusive breastfeeding / mixed or formula)	14 (64%) / 8 (36%)
Sex of child (boy / girl)	12 (45%) / 10 (55%)
Crying before breastfeeding (existence / absence) PD2 PD4	3 (14%) / 19 (86%) 8 (36%) / 14 (64%)
Data are shown as mean ± standard deviation. Values in parentheses are ranges or percentages
PD: postpartum day, STAI: State-Trait Anxiety Inventory

[insert Table 1 here]

All participants provided written informed consent after the purpose of the experiment was explained. The experimental protocol was in accordance with the Declaration of Helsinki (2013) and was approved by the Ethics Committee of the Nagasaki University Graduate School of Biomedical Sciences.

Procedures

As described previously [21], breastfeeding and sampling sessions were conducted by female experimenters (M.A-N and M.T) for approximately 60 min between 10:00 a.m. and 12:00 p.m. in a quiet, private, single room in the clinic. There was no music playing; the ceiling light was turned on, and the participant and experimenter were the only people in the room. Bathing for all newborns in the clinic was scheduled for 9:00 a.m. daily. We asked the participating mothers to abstain from breastfeeding after bathing and began the experiment at 10:00 a.m. in the private room. Participants did not feed the infants, nor were they swaddled during this time (9:00 a.m.–10:00 a.m.). The infant lay on a cot during the 30-min period before the experiment. Although some infants (PD2; n = 3/22, PD4; n = 8/22) were crying prior to breastfeeding (Table 1), there was no significant influence on OT when we compared OT between participants with and without infant crying using a t-test (P > 0.05). Participants were seated on comfortable chairs, and three samples of saliva were collected during the breastfeeding cycle. The first measurement was conducted 30 min before breastfeeding began (baseline), the second at the initiation of breastfeeding (+ 5 min after initiation of breastfeeding [feeding]), and the third was 30 min after the completion of breastfeeding (post-feeding). After breastfeeding was completed, most of the infants fell asleep, and the mothers placed them in the cot. We defined ΔOT (feeding minus baseline), which captures the OT level change in response to breastfeeding. As shown in Table 1, the third time point varied between individuals depending on the duration of breastfeeding. The experimenter did not terminate the breastfeeding session. For later correlation analyses, we calculated two indices, “Area under the curve with respect to increase” (AUCi) and “Area under the curve with respect to ground” (AUCg), which included the influence of individual duration of breastfeeding. The participants refrained from eating or drinking during the sample collection session.

Saliva sampling and storage

Saliva samples were collected from the participants using cotton Salivettes® (Sarstedt, Rommelsdorft, Germany) at all three time points. Participants were asked to place a Salivette® in their mouth and instructed to chew for 1 min until it was saturated with saliva. The Salivettes® were kept on ice for up to 2 h, before being centrifuged at 1,500 × g for 15 min at 4°C. Because of the short half-life of OT, the protease inhibitor aprotinin (500 KIU/mL, Sigma-Aldrich, St. Louis, MO, USA) was added to each collection tube to inhibit metabolic breakdown of the peptide. The liquid samples were stored at − 80°C.

Salivary OT measurement by the extracted method

OT concentrations were measured using an enzyme immunoassay kit (Enzo Life Sciences, Inc.). Briefly, saliva samples (500 µL per sample) were extracted using an Oasis PRiME HLB 96-well plate at 30 mg sorbent per well (SKU: 186008054, Waters Inc.) and evaporated at room temperature using compressed nitrogen. Each evaporated sample was reconstituted in 125 µL of assay buffer before OT measurement to provide a sufficient sample volume (100 µL per well). This practice ensured that the plated samples contained four-fold concentrated OT, which is sufficiently high to be read above the detection limit (15.0 pg/mL). The samples were assayed using a microplate reader (SPECTRA MAX 250, Molecular Devices, LLC.) for the 96-well format according to the manufacturer’s instructions. Because each saliva sample volume was less than 1.5–2 mL, we did not measure the samples using a duplicate assay. The mean intra- and inter-assay coefficients of variation calculated by the duplicated seven standard samples were 2.1% and 6.5%, respectively.

Salivary OT measurement by the unextracted method

To compare the differences between the extracted and unextracted method, we also measured the same samples using the unextracted method. Briefly, we used the lyophilization process instead of the extraction process to concentrate the samples four-fold overnight, reconstitute them in 125 µL of assay buffer, and store at − 20°C until they were assayed. The detailed procedures are described in our previous manuscript [21]. The mean intra- and inter-assay coefficients of variation calculated by the duplicated seven standard samples were 3.1% and 11.1%, respectively.

State and Trait Anxiety Inventory (STAI)

The STAI consists of 40 self-report items related to anxiety for both state (STAI-S) and trait (STAI-T) components, adapted and standardized for the Japanese population [24]. The STAI items were rated on a 4-point Likert scale. The range for each subtest (STAI-S and STAI-T) was 20–80, with a higher score indicating greater anxiety.

Statistical analysis

A one-way repeated-measures analysis of variance (ANOVA) was conducted to assess changes in OT levels across the breastfeeding cycle. Significant F ratios were further examined using post-hoc Fisher’s least significant difference test. To examine whether extracted and unextracted OT were correlated, Pearson’s correlation analyses were conducted for all the OT data; each time point (baseline, feeding, and post-feeding); and ΔOT, AUCi, and AUCg. To determine any associations between OT change indices (ΔOT, AUCi, and AUCg), clinical demographics, and STAI, Pearson’s correlation analyses or t-tests were conducted. All analyses were conducted using R (ver. 4.0.5) via the RStudio platform (ver. 1.2.5019) and the rstatix software package.

Salivary OT secretion patterns across the breastfeeding cycle measured by the extracted method

OT levels at baseline, feeding, and post-feeding are shown in Fig. 1 (left). One-way repeated-measures ANOVA revealed a significant difference in OT changes across the breastfeeding cycle at PD2 [F (2, 42) = 3.66; P < 0.05] but not at PD4 [F (2, 42) = 0.96; P = 0.39]. OT levels at PD2 increased significantly during feeding compared with baseline (t = 2.18, P = 0.03) and decreased between feeding and post-feeding, although this decrease was not statistically significant (t = -1.53, P = 0.13). OT levels after feeding were not significantly higher than those at baseline (t = 0.65, P = 0.52).

Salivary OT secretion patterns across the breastfeeding cycle measured by the unextracted method

OT levels at baseline, feeding, and post-feeding are shown in Fig. 1 (right). One-way repeated-measures ANOVA revealed a significant difference in OT changes across the breastfeeding cycle at PD2 [F (2, 42) = 5.43; P < 0.01] and PD4 [F (2, 42) = 4.91; P < 0.05]. OT levels at PD2 and 4 increased significantly during feeding compared with those at baseline (PD2: t = 2.69, P = 0.009, and PD4: t = 2.23, P = 0.03) and decreased between feeding and post-feeding (PD2: t = -2.02, P = 0.04), although this decrease was not statistically significant in the case of PD4 (t = -1.24, P = 0.22). OT levels after feeding were not significantly higher than those at baseline (PD2: t = 0.67, P = 0.50, and PD4: t = 0.99, P = 0.32).

Correlations between extracted and unextracted OT

As shown in Fig. 2, significant correlations were found between the extracted and unextracted OT at baseline (R = -0.38, P = 0.01) and ΔOT (R = 0.34, P = 0.02). No other pairwise correlations were significant (P > 0.05).

Correlations between OT change indices and clinical demographics and STAI

Pearson’s correlation coefficients between OT change indices (ΔOT, AUCi, and AUCg) and clinical demographics and STAI are shown in Additional Figure S1. Among the pairwise correlations, we found that all the OT change indices at PD2 measured by the extracted method were significantly negatively correlated with STAI-S at PD2 (PD2: ΔOT : R = -0.53, P = 0.01; AUCi: R = -0.53, P = 0.01; and AUCg: R = -0.45, P = 0.04) and had certain trends toward significance at PD4 (PD4: ΔOT : R = -0.40, P = 0.06, AUCi: R = -0.36, P = 0.10, and AUCg: R = -0.37, P = 0.09) (Fig. 3). In contrast to STAI-S, STAI-T was not correlated with the indices (ΔOT : R = 0.02, P = 0.92, AUCi: R = 0.007, P = 0.98, and AUCg: R = 0.04, P = 0.88). Although they were not correlated with the OT change indices at PD2, ΔOT, and AUCi at PD4 were negatively correlated with “Duration of labor” (ΔOT : R = -0.64, P = 0.001, and AUCi: R = -0.72, P = 0.0001) and “Induction medicine during labor” (ΔOT : R = -0.53, P = 0.01, and AUCi: R = -0.59, P = 0.004). “Duration of labor” and “Induction medicine during labor” were also well correlated with each other (R = 0.67, P = 0.0006); as induction medicine was practically used in cases with a long duration of labor, it can be interpreted to reflect the difficulty of labor. That is, patients with difficult labor would be less likely to show an OT increase by breastfeeding at PD4. In addition, “Induction medicine during labor” significantly attenuated the daily number of breastfeeding cycles performed on PD 1 (t = -2.25, P = 0.03) and 2 (t = -2.13, P = 0.046). Furthermore, we found that participants who were exclusively breastfeeding at the 1-month postpartum follow-up (n = 14) had only slightly significantly higher OT change indices at PD2, as measured by the extracted method, compared to those who were not exclusively breastfeeding (n = 8) (ΔOT : t = 1.84, P = 0.08, and AUCi: t = 1.69, P = 0.11).

In the present study, we investigated salivary OT changes following the breastfeeding cycle at PD2 and 4 in primiparous mothers who delivered vaginally and who had no abnormal symptoms during the postpartum period. We also utilized both extracted and unextracted methods for OT measurements and examined the correlation between the methods. The results showed that the extraction method showed a significant increase in OT concentration with breastfeeding at PD2, but not at PD4. The unextracted method showed a significant increase at both PD2 and 4. However, when we examined the correlation between the OT concentrations of the extracted and unextracted methods, we found a correlation between the baseline values and the inverse direction, but no correlations were found at the other time points or in the total data. However, the difference between breastfeeding and baseline was significantly positively correlated with each other. Hence, we subsequently dealt with OT data at PD2 measured via the extraction method to investigate whether increased OT associated with breastfeeding may decrease maternal anxiety. The results showed that the higher the OT change indices (ΔOT, AUCi, and AUCg), the lower the state anxiety at PD2 and 4; however, it was not associated with trait anxiety. There was also an association in that those who had higher OT change indices at PD2 were linked to the exclusive breastfeeding rate at the 1-month postpartum follow-up. Therefore, our results suggest that OT release by breastfeeding in the immediate postpartum period may reduce maternal anxiety and increase the rate of subsequent exclusive breastfeeding, as recommended by WHO/UNICEF [4].

The importance of the extraction method for OT measurement has been raised several times in the past [25–27]. Unlike our previous study [21] and other reports that used unextracted methods, the OT measurements in the present study were preprocessed using the extraction method. Moreover, we also measured the data using the same unextracted method as in the previous study and performed a novel comparison of the two conditions (the present unextracted data and population comprised a different dataset as that of our previous manuscript [21]). Thus, it can be observed that the true OT concentration, which is not a false positive, is clearly different from the absolute value of the unextracted method. The correlation of the baseline data alone was significant, but the direction of the correlation did not match. A number of publications [18–20, 28] have examined the relationship between baseline unextracted OT concentrations and various social and psychological metrics, although we believe that these need to be re-validated using extraction methods. In contrast, for ΔOT, there was a significant positive correlation between the extracted and unextracted methods. Even though false positives are included, the amount of change may still capture the overall change associated with the true OT change, even with the unextracted method. The following discussion is based on the results obtained from the extraction method.

Our results in the present study showed that breastfeeding increased salivary OT at PD2 but did not show a significant increase at PD4 or could not detect the changes by the saliva extraction method. Plasma OT, however, has been shown to capture an increase in OT even 2 months postpartum [29]. Therefore, this may be a phenomenon specific to salivary OT measurements. In particular, changes in salivary OT associated with breastfeeding appear to be influenced by the postpartum period. White-traut, et al. [19] measured salivary OT associated with the breastfeeding cycle within 8 months postpartum and found that baseline levels were the highest and decreased the most during breastfeeding. Carter, et al. (2007) [18] reported similar results. In addition, Jong, et al. (2015) [20] failed to observe changes in OT. Our PD4 results did not clearly decline, similar to those found in the studies by Carter and White-Traut; however, the results seemed to reflect a slight increase. We also examined the influences of other demographic and clinical factors that may inhibit OT secretion at PD4. There was an association between longer labor time and the use of induction medication, i.e., prolonged labor and a suppressed increase in OT at PD4. In addition, when examining the relationship between the use of induction medicines and the number of breastfeeding sessions 1–3 days postpartum, the use of induction medicines was associated with fewer breastfeeding sessions. Although positive feedback from OT is essential for parturition to instigate uterine contractions, it was thought that these participants may have been vulnerable to the neurobiological mechanism of OT release. For example, DNA methylation is one of the molecular mechanisms that suppresses gene expression [30]. Hiraoka et al. [31] reported that the more methylation in the OXT gene promoter region, the higher personal distress, an aspect of affective empathy of the mothers, and that there was brain gray matter volume reduction in the inferior temporal gyrus, which regulates empathy. Epigenetic mechanisms of the OXT gene may regulate OT secretion, leading to suppression of OT secretion during breastfeeding.

In the present study, for the first time, the changes in salivary OT with breastfeeding were captured using the extraction method. We showed that this OT increase was significantly associated with lower state anxiety, but it was not associated with trait anxiety. Unfortunately, we were unable to measure state anxiety as a time course change before and after breastfeeding. Although we only have the results of each one-time assessment at PD2 and 4, we were able to show an association between the increase in OT with breastfeeding and the decrease in state anxiety. However, we cannot deny the possibility that this relationship reflects an increase in OT because the original maternal anxiety was lower. Because there was no association between trait anxiety and OT and because the STAI was taken after the breastfeeding session, the lower maternal anxiety could be highly affected by the change in OT increase associated with breastfeeding. On the other hand, a higher OT increase with breastfeeding at PD2 was also causally associated with exclusive breastfeeding at the 1-month postpartum follow-up. WHO (1998) [4] encourages mothers to breastfeed for at least 6 months postpartum, recognizing physical and mental benefits to both the mother and infant. The present study results suggest that repeated self-exposure to endogenous OT triggered by breastfeeding in the immediate postpartum period in mothers could reduce their anxiety and lead to successful breastfeeding and ultimately to the health of both mother and child.

The present study had three major limitations. First, this study did not assess maternal anxiety using objective measures, such as behavioral tasks or brain MRI. In general, several studies have shown that OT has anxiolytic effects [32, 33], but whether self-exposure to endogenous OT caused by breastfeeding has anxiolytic effects has not been directly investigated. Although brain MRI would be difficult in the immediate postpartum period, it would have been beneficial if the anxiolytic effect could have been proven through measures, such as behavioral tasks. Second, this study was limited to primiparous women who delivered vaginally. A comprehensive study of the effects of wound pain and other factors in post-cesarean and multiparous women will be a future challenge. Third, the follow-up on breastfeeding rates was limited to 1 month. Although we showed that high OT secretion with immediate postpartum breastfeeding was associated with prolonged exclusive breastfeeding 1-month postpartum, it is desirable to clarify the longer-term effects using a prospective cohort design.

Breastfeeding in the early postpartum period resulted in an accompanying increase in endogenous OT, associated with lower maternal state anxiety, and potentially contributed to the exclusive breastfeeding rate at 1-month postpartum. This suggests that early postpartum breastfeeding could be the key to successful breastfeeding, as recommended by the WHO.

ANOVA, analysis of variance

AUCg, Area under the curve with respect to ground

AUCi, Area under the curve with respect to increase

OT, oxytocin

PD2, postpartum day 2

PD4, postpartum day 4

POMS, Profile of Mood States

STAI, State-Trait Anxiety Inventory

WHO, World Health Organization

Ethics approval and consent to participate

Consent for publication

Not applicable

Availability of data and materials

Data cannot be shared publicly because of the restrictions of the ethics committee. Data are available upon a reasonable request to the corresponding author for researchers who meet the criteria for access to confidential data.

Competing interests

The authors declare that they have no competing interests

Funding

All phases of this study were supported by JSPS KAKENHI Scientific Research (C) (25463477, 16K12096, 20K10834 to MN-A).

Authors' contributions

MN-A and SN designed the study. MN-A, MT, TT, TM, and SN conducted the experiment. MN-A, MT, HE, NS, WF, AM, KM, NH, and SN completed all analysis and prepared tables and figures. MN-A and SN wrote the main manuscript with critical revisions and supervision from HE, NS, WF, AM, KM and NH. All authors made substantial contributions to the analysis and data interpretation prior to agreeing upon key findings, conclusions, and approved the final manuscript.

Acknowledgments

We thank the staff at the Miyamura Obstetrics and Gynecology Clinic for their clerical support. We thank Mr. Hitoshi Takagi for lending us the draft chamber and space for the experiment in the Life Science Research Laboratory, University of Fukui.

Heinig MJ. Host defense benefits of breastfeeding for the Infant. Effect of breastfeeding duration and exclusivity. Pediatr Clin North Am. 2001;48:105 – 23, ix. doi: 10.1016/S0031-3955(05)70288-1.
Saarinen UM. Prolonged breast feeding as prophylaxis for recurrent otitis media. Acta Paediatr Scand. 1982;71:567–71. doi: 10.1111/j.1651-2227.1982.tb09476.x.
Howie PW, Forsyth JS, Ogston SA, Clark A, Florey CD. Protective effect of breast feeding against infection. BMJ. 1990;300:11–16. doi: 10.1136/bmj.300.6716.11.
Evidence for the ten steps to successful breastfeeding. World Health Organization; 1998.
Tharner A, Luijk MP, Raat H, Ijzendoorn MH, Bakermans-Kranenburg MJ, Moll HA, et al. Breastfeeding and its relation to maternal sensitivity and infant attachment. J Dev Behav Pediatr. 2012;33:396–404. doi: 10.1097/DBP.0b013e318257fac3.
Kim P, Feldman R, Mayes LC, Eicher V, Thompson N, Leckman JF, et al. Breastfeeding, brain activation to own infant cry, and maternal sensitivity. J Child Psychol Psychiatry. 2011;52:907–15. doi: 10.1111/j.1469-7610.2011.02406.x.
Kramer MS, Aboud F, Mironova E, Vanilovich I, Platt RW, Matush L, et al. Breastfeeding and child cognitive development: new evidence from a large randomized trial. Arch Gen Psychiatry. 2008;65:578–84. doi: 10.1001/archpsyc.65.5.578.
Feldman R, Bakermans-Kranenburg MJ. Oxytocin: a parenting hormone. Curr Opin Psychol. 2017;15: 13–18. doi: 10.1016/j.copsyc.2017.02.011.
Yamasue H, Okada T, Munesue T, Kuroda M, Fujioka T, Uno Y, et al. Effect of intranasal oxytocin on the core social symptoms of autism spectrum disorder: a randomized clinical trial. Mol Psychiatry. 2020;25:1849–58. doi: 10.1038/s41380-018-0097-2.
Uvnäs Moberg K, Ekström-Bergström A, Buckley S, Massarotti C, Pajalic Z, Luegmair K, et al. Maternal plasma levels of oxytocin during breastfeeding-A systematic review. PLOS ONE. 2020;15:e0235806. doi: 10.1371/journal.pone.0235806.
Weitzman RE, Leake RD, Rubin RT, Fisher DA. The effect of nursing on neurohypophyseal hormone and prolactin secretion in human subjects. J Clin Endocrinol Metab. 1980;51:836–39. doi: 10.1210/jcem-51-4-836.
McNeilly AS, Robinson IC, Houston MJ, Howie PW. Release of oxytocin and prolactin in response to suckling. Br Med J (Clin Res Ed). 1983;286:257–59. doi: 10.1136/bmj.286.6361.257.
Prime DK, Geddes DT, Hepworth AR, Trengove NJ, Hartmann PE. Comparison of the patterns of milk ejection during repeated breast expression sessions in women. Breastfeed Med. 2011;6:183–90. doi: 10.1089/bfm.2011.0014.
Grewen KM, Davenport RE, Light KC. An investigation of plasma and salivary oxytocin responses in breast- and formula-feeding mothers of infants. Psychophysiology. 2010;47:625–32. doi: 10.1111/j.1469-8986.2009.00968.x.
Feldman R, Gordon I, Zagoory-Sharon O. Maternal and paternal plasma, salivary, and urinary oxytocin and parent-infant synchrony: considering stress and affiliation components of human bonding. Dev Sci. 2011;14:752–61. doi: 10.1111/j.1467-7687.2010.01021.x.
Holt-Lunstad J, Birmingham W, Light KC. The influence of depressive symptomatology and perceived stress on plasma and salivary oxytocin before, during and after a support enhancement intervention. Psychoneuroendocrinology. 2011;36:1249–56. doi: 10.1016/j.psyneuen.2011.03.007.
Martin J, Kagerbauer SM, Gempt J, Podtschaske A, Hapfelmeier A, Schneider G. Oxytocin levels in saliva correlate better than plasma levels with concentrations in the cerebrospinal fluid of patients in neurocritical care. J Neuroendocrinol. 2018:e12596. doi: 10.1111/jne.12596.
Carter CS, Pournajafi-Nazarloo H, Kramer KM, Ziegler TE, White-Traut R, Bello D, et al. Oxytocin: behavioral associations and potential as a salivary biomarker. Ann N Y Acad Sci. 2007;1098:312–22. doi: 10.1196/annals.1384.006.
White-Traut R, Watanabe K, Pournajafi-Nazarloo H, Schwertz D, Bell A, Carter CS. Detection of salivary oxytocin levels in lactating women. Dev Psychobiol. 2009;51:367–73. doi: 10.1002/dev.20376.
Jong TR, Menon R, Bludau A, Grund T, Biermeier V, Klampfl SM, et al. Salivary oxytocin concentrations in response to running, sexual self-stimulation, breastfeeding and the TSST: the Regensburg Oxytocin Challenge (ROC) study. Psychoneuroendocrinology. 2015;62:381–88. doi: 10.1016/j.psyneuen.2015.08.027.
Niwayama R, Nishitani S, Takamura T, Shinohara K, Honda S, Miyamura T, et al. Oxytocin mediates a calming effect on postpartum mood in primiparous mothers. Breastfeed Med. 2017;12:103–9. doi: 10.1089/bfm.2016.0052.
McNair DMLM, Droppleman LF. Profile of Mood States. Manual. Educational and industrial testing service, CA; 1971.
Zonana J, Gorman JM. The neurobiology of postpartum depression. CNS Spectr. 2005;10:792–9, 805. doi: 10.1017/s1092852900010312.
Nakazato K, Mizuguti T. Development and validation of Japanease version of State-Trait Anxiety Inventory: A study with female subjects. Jap Soc Psychosom Med. 1982;22:107–12.
Saxbe D, Khaled M, Horton KT, Mendez AJ. Maternal prenatal plasma oxytocin is positively associated with prenatal psychological symptoms, but method of immunoassay extraction may affect results. Biol Psychol. 2019;147:107718. doi: 10.1016/j.biopsycho.2019.107718.
Morris AR, Turner A, Gilbertson CH, Corner G, Mendez AJ, Saxbe DE. Physical touch during father-infant interactions is associated with paternal oxytocin levels. Infant Behav Dev. 2021;64:101613. doi: 10.1016/j.infbeh.2021.101613.
Szeto A, McCabe PM, Nation DA, Tabak BA, Rossetti MA, McCullough ME, et al. Evaluation of enzyme immunoassay and radioimmunoassay methods for the measurement of plasma oxytocin. Psychosom Med. 2011;73:393–400. doi: 10.1097/PSY.0b013e31821df0c2.
Chu C, Hammock EAD, Joiner TE. Unextracted plasma oxytocin levels decrease following in-laboratory social exclusion in young adults with a suicide attempt history. J Psychiatr Res. 2020;121:173–81. doi: 10.1016/j.jpsychires.2019.11.015.
Whitley J, Wouk K, Bauer AE, Grewen K, Gottfredson NC, Meltzer-Brody S, et al. Oxytocin during breastfeeding and maternal mood symptoms. Psychoneuroendocrinology. 2020;113:104581. doi: 10.1016/j.psyneuen.2019.104581.
Nishitani S, Parets SE, Haas BW, Smith AK. DNA methylation analysis from saliva samples for epidemiological studies. Epigenetics. 2018;13:352–62. doi: 10.1080/15592294.2018.1461295.
Hiraoka D, Nishitani S, Shimada K, Kasaba R, Fujisawa TX, Tomoda A. Epigenetic modification of the oxytocin gene is associated with gray matter volume and trait empathy in mothers. Psychoneuroendocrinology. 2021;123:105026. doi: 10.1016/j.psyneuen.2020.105026.
Yoon S, Kim YK. The role of the oxytocin system in anxiety disorders. Adv Exp Med Biol. 2020;1191:103–20. doi: 10.1007/978-981-32-9705-0_7.
Naja WJ, Aoun MP. Oxytocin and anxiety disorders: translational and therapeutic aspects. Curr Psychiatry Rep. 2017;19:67. doi: 10.1007/s11920-017-0819-1.

No competing interests reported.

092821SupplementaryInformation.docx

Download PDF

Editorial decision: Major revision
03 Jun, 2022
Reviews received at journal
30 May, 2022
Reviewers agreed at journal
19 May, 2022
Reviewers invited by journal
19 May, 2022
Editor assigned by journal
19 May, 2022
Editor invited by journal
12 Mar, 2022
Submission checks completed at journal
12 Mar, 2022
First submitted to journal
05 Mar, 2022

You are reading this latest preprint version

Endogenous oxytocin levels in extracted saliva elevates during breastfeeding correlated with lower postpartum anxiety in primiparous mothers

Status:

Version 1

Abstract

Background

Methods

Results

Conclusions

Figures

Background

Methods

Participants

Procedures

Saliva sampling and storage

Salivary OT measurement by the extracted method

Salivary OT measurement by the unextracted method

State and Trait Anxiety Inventory (STAI)

Statistical analysis

Results

Salivary OT secretion patterns across the breastfeeding cycle measured by the extracted method

Salivary OT secretion patterns across the breastfeeding cycle measured by the unextracted method

Correlations between extracted and unextracted OT

Correlations between OT change indices and clinical demographics and STAI

Discussion

Conclusion

Abbreviations

Declarations

References

Additional Declarations

Supplementary Files

Status:

Version 1