A Comparison of Breast Milk and Sucrose in Reducing Neonatal Pain During Eye Exam for Retinopathy of Prematurity

Abstract

Background:

This double-blind randomized controlled experimental study aimed to determine the effects of breast milk and sucrose in reducing pain in preterm infants during retinopathy of prematurity (ROP) examination.

Materials and Methods:

This study was conducted with 60 preterm infants (breast milk group = 20, sucrose group = 20, and control/distilled water group = 20) meeting the inclusion criteria. The data were collected with the use of an Infant Evaluation Form, Procedure Monitoring Form, and Premature Infant Pain Profile (PIPP). The preterm infants were provided with 1 mL of breast milk, sucrose, and distilled water before the ROP examination. The pain level in preterm infants was measured by the PIPP 5 minutes before, during, and 5 minutes after the ROP examination. The ROP examinations were video recorded, and videos were evaluated by three observers blinded to the study.

Results:

No significant difference was determined between the three groups in terms of their postconceptional and postnatal ages, their body weights at birth and during the ROP examination. The PIPP scores of the preterm infants in the three groups were higher during the ROP examination and were not significantly different. The PIPP scores of the control group were significantly higher than those in the breast milk and sucrose groups after the ROP examination (p < 0.001). The preterm infants in the breast milk group recovered and returned to their initial values more quickly after the ROP examination than the infants in the sucrose group.

Conclusion:

To reduce pain in preterm infants during ROP examination, breast milk is recommended.

Introduction

The technological developments in Neonatal Intensive Care Units (NICUs) and the rapid development in the knowledge and treatment opportunities related to the care of preterm infants have increased the survival of newborns at risk.^1,2 The decreased mortality in preterm infants has led to increased morbidity, requiring long-term hospitalization and frequent follow-ups. These infants experience pain and stress resulting not only from the disease processes, but also the great number of invasive procedures during their hospitalization in the NICU.^3,4 One of these painful procedures for preterm infants is retinopathy of prematurity (ROP) examination. Untreated ROP may lead to total vision loss or myopia, strabismus, amblyopia, macular ectopia, retinal detachment, angle closure glaucoma, and pupillary block.^5–8 Consequently, the American Academy of Pediatrics, American Academy of Ophthalmology, and American Association for Pediatric Ophthalmology and Strabismus have issued a joint statement on the importance of routine ROP examinations for preterm infants with a birth weight of <1,500 g or a gestational age of 32 weeks or less.^9,10

The ROP examination involves the application of dilating drops and binocular indirect ophthalmoscopy (BIO) with a lid speculum and scleral depression. The examination procedure, which includes fixation of the infant's head and opening of the eyelids, often using an eye speculum and a light beam, results in signs that are associated with pain, discomfort, and stress.^9,11–14 The International Evidence-Based Group for Neonatal Pain put the ROP examination on the list of painful procedures performed in the NICU.¹⁵ Systemic complications, including an increase in blood pressure, heart rate, respiratory rate, oxygen requirement to correct increased oxygen desaturation, and serum cortisol levels during and after the ROP examination, have been reported.^{5,12–14,16–25} In an attempt to reduce the pain and discomfort associated with the ROP examination, various nonpharmacological methods have been used, including giving nipples, swaddling, kangaroo care, music therapy, oral sucrose in single or repetitive doses, breastfeeding, and individualized developmental care practices,^26–35 though most studies have not compared these various techniques. This double-blind randomized controlled experimental study aimed to determine the comparative effects of breast milk and sucrose in reducing the pain of preterm infants during the ROP examination.

Materials and Methods

Sampling

This study was initiated with 63 preterm infants; however, 3 preterm infants (2 in the control group, 1 in the breast milk group) were excluded due to a poor quality video recording of the ROP examination. In total, 60 preterm infants meeting the inclusion criteria (breast milk group = 20, sucrose group = 20, and control/distilled water group = 20) during the period from August 10, 2010 to March 10, 2011 were included. Ethics committee approval (Date: August 5, 2010, No: 2010/87), the permission of the institution (Date: July 23, 2010, No: 3163), and a written consent form were obtained from the parents of the preterm infants before the study. Power analysis performed by a professional statistician (Associate Professor PhD in Department of Biostatistics and Medical Informatics, Faculty of Medicine) was used to calculate the sample size, which was detected as 0.97 after the study.

Inclusion criteria

Preterm infants with a birth weight ≤1,500 g and gestational age of ≤32 weeks.

Exclusion criteria

Preterm infants diagnosed with a congenital anomaly, hydrocephalus, necrotizan enterocolitis, indirect hyperbilirubinemia, and receiving ventilatory support or analgesic drug treatment.

Clinical data were collected with the use of an Infant Evaluation Form and a Procedure Monitoring Form; the pain of the preterm infants was assessed via the Premature Infant Pain Profile (PIPP).

Infant evaluation form

The characteristics of the infants were recorded.

The PIPP was used to measure pain and discomfort. The PIPP is a scale to diagnose pain developed by Stevens et al.³⁶ for preterm infants. Its validity and reliability in the Turkish population was tested by Derebent in 2006.³⁷ The scale consists of questions with seven factors: gestational age, behavioral status, the highest heart rate value, the lowest oxygen saturation value, wrinkling of the forehead, squinting, and expansion of nose wings. The lowest and highest scores of this scale are 0 and 21, respectively; the scale assesses the pain of the infants over the total score.^36,37

Study design

The assignment to groups of the preterm infants meeting the inclusion criteria was performed by a nurse employed in the service, away from the researchers. The preterm infants were randomly assigned to the breast milk, sucrose, and control groups.

The eyes of the preterm infants were dilated 1 hour before the examination by dropping 0.5% cyclopentolate and 2.5% phenylephrine into their eyes—one drop for each eye at 5-minute intervals, three times in total. The preterm infants were fed and their diapers were changed at least 30 minutes before the examination. No painful procedure was applied to the preterm infants at least 30 minutes prior and 15 minutes after the ROP examination. The ROP examinations were performed in a separate place in the clinic, in a cot to facilitate video recording. One of the researchers recorded the physiological measurements (heart rate, respiratory rate, and oxygen saturation) of the preterm infants on the Procedure Monitoring Form and scored the PIPP 5 minutes before and 5 minutes after the ROP examination. Before the ROP examination (2 minutes), Group 1 was provided with 1 mL breast milk; Group 2 was provided with 1 mL 33% sucrose; and Group 3 was provided with 1 mL distilled water via nipples+injector. Another researcher video recorded the ROP examinations, which lasted on average 2.33 ± 0.85 minutes. The behavioral responses of the infants during the ROP examination and physiological measurements were assessed using these video records.

The ROP examinations were carried out by the same ophthalmologist. At the beginning of the examination, first the presegment and then the fundus were examined using a scleral depressor, binocular indirect ophthalmoscope, and 20-diopter lenses upon placing the eyelid speculum after 0.5% proparacaine HCl had been dropped as the topical anesthetic. In this double-blind study, only the researcher providing breast milk, sucrose, or distilled water to the preterm infants before the examination (2 minutes) knew the group assignment of the preterm infants. Hence, this researcher performed the video recordings. The researcher carrying out the physiological measurements 5 minutes before and 5 minutes after the examination also performed the video recordings without having any information on the group assignment of the preterm infants.

Analysis of the video records of the ROP examination

Three independent specialists (two nurses specialized in pediatric nursing and a neonatologist who had no information about each other analyzed the video records of the ROP examination and scored the PIPP. Inter-observer consistency analysis was performed for each item of the PIPP after the specialists had assessed the pain and determined the PIPP scores of all the preterm infants, which ranged between 0.90 and 1.00. The PIPP scores submitted by the three specialists were added and averaged by the researcher after the inter-observer consistency had been ensured and these averages were used. The average duration of the ROP examination was 2.33 ± 0.85 minutes.

Data analysis

The data were analyzed using the IBM SPSS Statistics 21.0 (Chicago, IL) package. In addition, descriptive statistics (percentage, arithmetic mean, standard deviation, and median), Shapiro–Wilk test, Chi-square test, analysis of variance (ANOVA), and Kruskal–Wallis ANOVA were used in the study along with the two-way ANOVA and Student-Newman-Keuls Method for multiple repetitive measurements. The intra-class correlation coefficient (ICC) was used for the inter-observer consistency among the three specialists assessing the video records of the ROP examination. p < 0.05 was accepted as being statistically significant.

Results

The majority of the preterm infants included in the study had a gestational age of >28 weeks, birth weight of >1,000 g, and body weight of >1,500 g during the ROP examination; were female and orally fed; and there was no statistically significant difference between the groups (Table 1).

Table 1.

The Characteristics of the Preterm Infants

	Breast milk group (n = 20)		Sucrose group (n = 20)		Control group (n = 20)
Characteristics	n	%	n	%	n	%	χ²	p
Gender
Male	9	45.0	6	30.0	12	60.0	3.636	0.162
Female	11	55.0	14	70.0	8	40.0
Gestational age (weeks)
≤27.6	1	5.0	5	25.0	7	35.0	5.719^*	0.058
>28	19	95.0	15	75.0	13	65.0
Postconceptional age (weeks)
≤36.6	12	60.0	16	80.0	15	75.0	2.043^*	0.450
>37	8	40.0	4	20.0	5	25.0
Birth weight (g)
≤1,000	2	10.0	7	35.0	9	45.0	6.401^*	0.042
>1,000	18	90.0	13	65.0	11	55.0
Body weight (g)
≤1,500	5	25.0	2	10.0	4	20.0	1.586^*	0.589
>1,500	15	75.0	18	90.0	16	80.0
Feeding style
Orally	17	85.0	18	90.0	14	70.0	2.648^*	0.339
Orally+NG	3	15.0	2	10.0	6	30.0

Fisher's exact test.

No significant difference was detected between the breast milk, sucrose, and control groups in terms of mean scores for heart rate and oxygen saturation (p = 0.256, p = 0.763, respectively). As expected, a significant difference in response to the procedures was detected between all the heart rate and oxygen saturation measurements (before, during, and after) of the preterm infants in the three groups (p < 0.001, p < 0.001, respectively). The group and time interaction of the mean scores for heart rate and oxygen saturation were not significant (p = 0.622, p = 0.055, respectively) (Table 2).

Table 2.

Heart Rate and Oxygen Saturation Measurements of the Preterm Infants Before, During, and After Retinopathy of Prematurity Examination

		Heart rate		Oxygen saturation
Groups	Time	Mean ± SD	Median (Min.–Max.)	Mean ± SD	Median (Min.–Max.)
Breast milk group (n = 20)	Before the examination	151.80 ± 11.76	152.0 (130–174)	91.10 ± 5.48	97.0 (80–100)
	During the examination	176.59 ± 18.18	176.33 (140–202)	89.88 ± 7.72	91.20 (76.77–100)
	After the examination	159.70 ± 12.86	160.0 (128–180)	91.05 ± 7.88	93.0 (75–100)
Sucrose group (n = 20)	Before the examination	155.20 ± 16.72	159.0 (116–188)	91.55 ± 5.16	92.0 (80–100)
	During the examination	183.42 ± 21.71	184.13 (145.77–230)	88.93 ± 6.57	90.11 (73.38–95.5)
	After the examination	165.70 ± 9.99	164.0 (148–192)	88.30 ± 8.73	89.5 (72–100)
Control group (n = 20)	Before the examination	153.90 ± 6.23	156.0 (140–166)	93.10 ± 5.06	93.5 (85–100)
	During the examination	174.94 ± 12.41	173.36 (152.92–202.22)	90.00 ± 5.07	90.97 (78.75–98.15)
	After the examination	160.50 ± 9.40	158.0 (148–186)	90.25 ± 5.81	92.0 (81–98)

		F	P	F	p
Test	Time	94.668	<0.001	19.570	<0.001
	Time+Group	0.658	0.622	2.388	0.055
	Group	1.396	0.256	0.271	0.763

SD, standard deviation.

The mean PIPP scores of the breast milk, sucrose, and control groups before and during the ROP examination were similar (p > 0.05) (Table 3). The mean PIPP scores after the ROP examination were not significantly different between the breast milk and sucrose groups, however, there was a significant difference between the control group and the breast milk and sucrose groups (p < 0.001). This significant difference was determined to be due to the higher mean PIPP scores of the preterm infants in the control group after the ROP examination (p < 0.05) (Table 3). Breast milk feeding resulted in the infants returning to the initial values more quickly. After the ROP examination, the PIPP scores were 3.20 in the breast milk group and 3.65 in the sucrose group. Also, the mean heart rates were lower and the mean oxygen saturation rates were higher in the breast milk group than in the sucrose group and these values were similar to those before the ROP examination (Tables 2 and 3).

Table 3.

The Premature Infant Pain Profile Scores of the Preterm Infants Before, During, and After Retinopathy of Prematurity Examination

		PIPP scores
Groups	Time	Mean ± SD	Median (Min.–Max.)
Breast milk group (n = 20)	Before the examination	2.10 ± 0.55	2.0 (1.0–4.0)
	During the examination	16.06 ± 2.41	16.0 (7.0–19.0)
	After the examination	3.20 ± 1.36	3.0 (1.0–6.0)
Sucrose group (n = 20)	Before the examination	2.05 ± 0.75	2.0 (1.0–3.0)
	During the examination	15.58 ± 2.02	15.66 (10.0–18.67)
	After the examination	3.65 ± 1.78	3.50 (1.0–7.0)
Control group (n = 20)	Before the examination	2.50 ± 0.68	2.0 (2.0–4.0)
	During the examination	15.88 ± 1.66	16.50 (12.0–18.33)
	After the examination	5.90 ± 1.61	6.0 (2.0–8.0)

		F	p
Test	Time	1,682.547	<0.001
	Time+Group	6.434	<0.001
	Group	5.742	0.005

PIPP, Premature Infant Pain Profile; SD, standard deviation.

Discussion

Although the ROP examination was performed with the use of topical anesthetic drops, studies have demonstrated that infants experience a lot of pain during the examination.^{5,13,14,17,33–35} In the current literature, a few studies were conducted aimed at reducing pain during the ROP examination by implementing nonpharmacological methods (including giving nipples, swaddling, providing sucrose in single or repetitive doses, and individualized developmental care practices) that have existed since the last decade.^{5,17–19,33–35}

Breast milk, being safe, effective, and natural, provides an analgesic effect at no cost.³⁸ Breast milk contains a higher concentration of tryptophan, a precursor of both serotonin and melatonin. Melatonin has been shown to increase the concentration of beta-endorphins and so could be one of the mechanisms for the nociceptive effects of breast milk.^39,40 Serotonin, a neurotransmitter synthesized from tryptophan, regulates cognition, attention, emotion, pain, sleep, and arousal.^40,41 The analgesic effect of breast milk was preferred to the induction of opioids due to its content of lipids, proteins, and other tastes, and because it blocks the pain fibers reaching the spinal cord with the inhibition of pain transmission.⁴² Studies have revealed that breast milk, either through breastfeeding or nipples, is an effective strategy for reducing pain.^43–45

To the best of our knowledge, there are few number of studies^34,35 providing breast milk to reduce pain during the ROP examination in the current literature and our current study is the third one in this area of research. In the current study, all the mean heart rate measurements (before, during, and after the examination) of the preterm infants in the three groups were similar, and they were also higher during the ROP examination. This may be due to the long and painful nature of the ROP examination and the preterm infants continuously cried in trying to cope with this experience of pain during the examination. The studies on the pain of preterm infants during the ROP examination also referred to the higher heart rates of infants during the examination.^5,18,33 There was no significant difference between the mean oxygen saturation measurements (before, during, and after the examination) of the preterm infants in the three groups. The mean oxygen saturation measurements were lower during the ROP examination (p < 0.001) (Table 2). Other studies also reported a decrease in the oxygen saturation of preterm infants during the ROP examination.^{5,11,17,18,20,33,46} Laws et al.¹¹ analyzed the ROP imaging process and determined that the physical manipulation of the eyes caused serious changes in oxygen saturation, blood pressure, and heart rate. Similarly, Rush et al.³³ detected no difference between the experimental and control groups in terms of the changes in the time of crying, heart and respiratory rates, and the oxygen saturation of preterm infants, during the ROP examination in their study which provided 24% sucrose.

The mean PIPP scores of the preterm infants in the three groups during the examination were higher (Table 3) (p < 0.001). This may be the result of the long and painful nature of the ROP examination, despite the use of topical anesthetic eye drops. Also, placing a speculum on the eyes, putting pressure on the eyeball, and the brightness of the ophthalmoscope may have additional negative effects on preterm infants in terms of experiencing more pain.^11–14 Grabska et al.¹⁸ also determined the highest pain perception in the sucrose and control groups during the ROP examination despite the use of topical anesthetic eye drops in their study, which provided a single dose of sucrose. They found no difference in the heart and respiratory rates, blood pressure, and the PIPP scores between the two groups during the retinal examination. Mitchell et al.¹⁷ found that infants administered sucrose in repetitive doses during the ROP examination had a lower pain perception. Olsson and Eriksson⁴⁷ evaluated the use of oral glucose versus sterile water during the examination and detected no difference between the groups for PIPP scores, heart rate changes, or duration of crying.

In the current literature, some studies compared breast milk and sucrose.^34,35,48,49 Simonse et al.⁴⁸ found no difference between breast milk and sucrose in reducing the pain of preterm infants during the heel lance procedure. Ribeiro et al.³⁵ evaluated the use of breast milk versus sucrose for pain relief during retinal eye examinations and there was no significant difference between groups for crying, heart rate, and salivary cortisol levels. Our results are in accordance with their study. Rosali et al.³⁴ found breast milk to be effective in reducing the pain of preterm infants during the ROP examination and Gal et al.⁴⁹ found that 24% sucrose had the same effect. However, we found that both breast milk and sucrose were ineffective in reducing the pain of preterm infants during the ROP examination.

In the current study, the mean PIPP scores after the ROP examination were not significant for the breast milk and sucrose groups, however, there was a significant difference for the control and the breast milk and sucrose groups (p < 0.001). After the ROP examination, it can be concluded that breast milk and sucrose are effective in reducing pain among preterm infants. This may be due to breast milk and sucrose, two nonpharmacological methods, relieving preterm infants and enabling them to recover and reach their initial values more quickly.

The preterm infants in the breast milk group recovered and returned to their initial values more quickly after the ROP exam than the infants in the sucrose group (Table 3). This may be interpreted as a quicker recovery and less painful experience for the infants after the ROP examination. Also, the mean heart rates were lower and the mean oxygen saturation rates were higher in the breast milk group than in the sucrose group and these values were similar to those before the ROP examination.

The lower mean PIPP scores of the breast milk group after the ROP examination than those of the other two groups may be interpreted as a quicker recovery and less painful experience for the infants after the ROP examination. Similarly, Rosali et al.³⁴ found breast milk to be more efficient at decreasing pain and helping in reaching the initial values after the ROP examination. In the current study, breast milk was superior to sucrose in terms of reducing pain, as the long duration of the ROP examination and the effect of sucrose on pain might have decreased 5 minutes after the ROP examination.

Conclusion

In conclusion, based on the finding that the breast milk group recovered and reached their initial physiological and behavioral values more quickly after the ROP examination, and due to the superiority of breast milk as being a safe, effective, available, and natural analgesic at no cost, breast milk is better than sucrose during the ROP examination. It is recommended that similar studies should be conducted providing repetitive doses of breast milk and sucrose due to the long and painful nature of the ROP examination.

Footnotes

Acknowledgments

We would like to thank Ferhan Elmali (Associate Professor, PhD, in Department of Biostatistics and Medical Informatics, Faculty of Medicine, İzmir Katip Çelebi University) for statistical analysis. This article has received funding from Scientific Research Project Coordination Unit Erciyes University (Project Code: TSY-10-3381).

Disclosure Statement

No competing financial interests exist.

References

Ballot

, Chirwa

, Cooper

. Determinants of survival in very low birth weight neonates in a public sector hospital in Johannesburg. BMC Pediatr, 2010; 10:10–11.

Daga

, Daga

, Mhatre

, et al. Enhancing neonatal survival: What can we do today?. J Perinatol, 2016; 36:681–684.

Carbajal

, Rousset

, Danan

, et al. Epidemiology and treatment of painful procedures in neonates in intensive care units. JAMA, 2008; 300:60–70.

Roofthooft

DWE

, Simons

SHP

, Anand

KJS

, et al. Eight years later, are we still hurting newborn infants?. Neonatology, 2014; 105:218–226.

Boyle

, Freer

, Khan-Orakzai

, et al. Sucrose and non-nutritive sucking for the relief of pain in screening for retinopathy of prematurity: A randomised controlled trial. Arch Dis Child Fetal Neonatal Ed, 2006; 91:166–168.

Samra

, McGrath

. Pain management during retinopathy of prematurity eye examinations. Adv Neonatal Care, 2009; 9:99–110.

Tolsma

, Allred

, Chen

, et al. Neonatal bacteremia and rethinopathy of prematurity. The ELGAN study. Arch Ophthalmol, 2011; 129:1555–1563.

Özkan

, Köksal

. Retinopathy of prematurity. J Curr Pediatr, 2005; 2:24–28.

Upadhyay

, Kumar Gupta

. Retinopathy of prematurity (ROP) and its associated pain. Indian J Pediatr, 2015; 82:673–674.

10.

American Academy of Ophthalmology, American Association for Pediatric Ophthalmology and Strabismus, American Academy of Pediatrics Section on Ophthalmology. Screening examination of premature infants for retinopathy of prematurity. Pediatrics, 2006; 117:572–576.

11.

Laws

, Morton

, Weindling

, et al. Systemic effects of screening for retinopathy of prematurity. Br J Ophthalmol, 1996; 80:425–428.

12.

Haigh

, Chiswick

, O'Donoghue

, et al. Retinopathy of prematurity: Systemic complications associated with different anaesthetic techniques at treatment. Br J Ophthalmol, 1997; 81:283–287.

13.

Belda

, Pallas

, Cruz

, et al. Screening for retinopathy of prematurity: Is it painful?. Biol Neonate, 2004; 86:195–200.

14.

Kirchner

, Jeitler

, Pollak

, et al. Must screening examinations for retinopathy of prematurity necessarily be painful?. Retina, 2009; 29:586–591.

15.

Anand

KJS

. International Evidence-based Group for Neonatal Pain. Consensus statement for the prevention and management of pain in the newborn. Arch Pediatr Adolesc Med, 2001; 155:173–180.

16.

Hartrey

. Anaesthesia for the laser treatment of neonates with retinopathy of prematurity. Eye, 2007; 21:1025–1027.

17.

Mitchell

, Stevens

, Mungan

, et al. Analgesic effects of oral sucrose and pacifier during eye examinations for retinopathy of prematurity. Pain Manag Nurs, 2004; 5:160–168.

18.

Grabska

, Walden

, Lerer

, et al. Can oral sucrose reduce the pain and distress associated with screening for retinopathy of prematurity?. J Perinatol, 2005; 25:33–35.

19.

Kleberg

, Warren

, Norman

, et al. Lower stress responses after newborn individualized development care and assessment program care during eye screening examinations for retinopathy of prematurity: A randomized study. Pediatrics, 2008; 121:1267–1278.

20.

Mehta

, Adams

, Bunce

, et al. Pilot study of the systemic effects of three different screening methods used for retinopathy of prematurity. Early Hum Dev, 2005; 8:355–360.

21.

Kandasamy

, Smith

, Wright

, et al. Pain relief for prematüre infants during ophthalmology assessment. J AAPOS, 2011; 15:276–280.

22.

Slevin

, Murphy

, Daly

, et al. Retinopathy of prematurity screening, stress related responses, the role of nesting. Br J Ophthalmol, 1997; 81:762–764.

23.

Mitchell

, Green

, Jeffs

, et al. Physiologic effects of retinopathy of prematurity screening examinations. Adv Neonatal Care, 2011; 11:291–297.

24.

Rush

, Rush

, Nicolau

, et al. Systemic manifestations in response to mydriasis and physical examination during screening for retinopathy of prematurity. Retina, 2004; 24:242–245.

25.

Wood

, Kaufman

. Apnea and bradycardia in two premature infants during routine outpatient retinopathy of prematurity screening. J AAPOS, 2009; 13:501–503.

26.

Seo

, Lee

, Ahn

. Effects of kangaroo care on neonatal pain in South Korea. J Trop Pediatr, 2016; 62:246–249.

27.

Leng

, Zheng

, Zhang

, et al. Combined non-pharmacological interventions for newborn pain relief in two degrees of pain procedures: A randomized clinical trial. Eur J Pain, 2016; 20:989–997.

28.

Pillai Riddell

, Racine

, Gennis

, et al. Non-pharmacological management of infant and young child procedural pain. Cochrane Database Syst Rev, 2015; 2:CD006275.

29.

Obeidat

, Shuriquie

. Effect of breast-feeding and maternal holding in relieving painful responses in full-term neonates: A randomized clinical trial. J Perinat Neonatal Nurs, 2015; 29:248–254.

30.

Azarmnejad

, Sarhangi

, Javadi

, et al. The effect of mother's voice on arterial blood sampling induced pain in neonates hospitalized in neonate intensive care unit. Glob J Health Sci, 2015; 7:198–204.

31.

Potana

, Dongara

, Nimbalkar

, et al. Oral sucrose for pain in neonates during echocardiography: A randomized controlled trial. Indian Pediatr, 2015; 52:493–497.

32.

Gao

, Xu

, Gao

, et al. Effect of repeated kangaroo mother care on repeated procedural pain in preterm infants: A randomized controlled trial. Int J Nurs Stud, 2015; 52:1157–1165.

33.

Rush

, Rush

, Ighanı

, et al. The effects of comfort care on the pain response in preterm infants undergoing screening for retinopathy of prematurity. Retina, 2005; 25:59–62.

34.

Rosali

, Nesargi

, Mathew

, et al. Efficacy of expressed breast milk in reducing pain during rop screening-a randomized controlled trial. J Trop Pediatr, 2015; 61:135–138.

35.

Ribeiro

, Castral

, Montanholi

, et al. Human milk for neonatal pain relief during ophthalmoscopy. Rev Esc Enferm USP, 2013; 47:1039–1045.

36.

Stevens

, Johnston

, Petryshen

, et al. Premature infant pain profile: Development and initial validation. Clin J Pain, 1996; 12:13–22.

37.

Derebent

, Yiğit

. Turkish reliability and validity study of Premature Infant Pain Profile. Firat Univ Health Sci Med J, 2015; 29:97–102

38.

Reece-Stremtan

, Gray

, The Academy of Breastfeeding Medicine. Nonpharmacological management of procedure-related pain in the breastfeeding infant, revised 2016. Breastfeed Med, 2016; 11:1–5.

39.

Barrett

, Kent

, Voudoris

. Does melatonin modulate beta-endorphin, corticosterone, and pain threshold?. Life Sci, 2000; 66:467–476.

40.

Rehmani

, Farhan

, Hadi

. DNA binding and its degradation by the neurotransmitter serotonin and its structural analogues melatonin and tryptophan: Putative neurotoxic mechanism. J Mol Genet Med, 2016; 10:223.

41.

Brummelte

, Mc Glanaghy

, Bonnin

, et al. Developmental changes in serotonin signaling: Implications for early brain functıon, behavior and adaptation. Neuroscience, 2017; 342:212–231.

42.

Gray

, Miller

, Philipp

, et al. Breastfeeding isanalgesic in healthy newborns. Pediatrics, 2002; 109:590–593.

43.

Upadhay

, Aggarwal

, Narayan

, et al. Analgesic effect of expressed breastmilk in proceduralpain in termneonates: A randomized, placebo-controlled, doubleblind trial. Acta Pediatr, 2004; 93:518–522.

44.

Codipietro

, Ceccarelli

, Ponzone

. Breastfeeding or oral sucrose solution in term neonates receiving heel lance: A randomized, controlled trial. Pediatrics, 2008; 122:e716–e720.

45.

Shah

, Herbozo

, Aliwalas

, et al. Breastfeeding or breastmilk for procedural pain in neonates. Cochrane Database Syst Rev, 2012; 12:CD004950.

46.

Clarke

, Hodges

, Noel

, et al. The oculocardiac reflex during ophthalmoscopy in premature infants. Am J Ophthamol, 1985; 99:649–651.

47.

Olsson

, Eriksson

. Oral glucose for pain relief during eye examinations for retinopathy of prematurity. J Clin Nurs, 2011; 20:1054–1059.

48.

Simonse

, Mulder

GHP

, Van Beek

. Analgesic effect of breast milk versus sucrose for analgesia during heel lance in late preterm infants. Pediatrics, 2012; 129:657–663.

49.

Gal

, Kissling

, Young

, et al. Efficacy of sucrose to reduce pain in premature infants during eye examinations for retinopathy of prematurity. Ann Pharmacother, 2005; 39:1029–1033.