PYK Wu

Although light has been used as an indicator of hemoglobin oxygenation for almost half a century, the concept of using light in the near-infrared (NIR) range of the spectrum to monitor tissue oxygenation and hemodynamics was first described by Jobsis¹ and his colleagues in 1977.² Since then the technology has been applied to provide information of hemodynamic changes and metabolic status in intact tissues and body parts in animal research, adults and infants in various states of health. Of particular interest to neonatologist is the pioneer work of Brazy and her colleagues in applying the near-infrared (NIRS) method to obtain information on the intraneuronal availability of oxygen and redox state of intraneuronal enzyme cytochrome aa₃ (Cyt aa₃), that catalyzes the use of oxygen in the process of oxidative phosphorylation, in neonates with hypoxia, hypertensive or crying episodes, and during various procedures such as temperature taking, echocardiography, and gavage feeding.^3-5 Concurrently and since then a large number of investigators have contributed many valuable information with the use of NIRS in neonatal medicine. This review is focused on summarizing some of the major applications of this technique in neonates.

Near-infrared spectroscopy method is based on the fact that light in the near-infrared (NIR) range (within 700-1000 nm) can pass through skin, bone and other tissues relatively easily and that there are characteristic absorption bands of oxygenated and deoxygenated hemoglobin (HbO₂ and Hb-) and of Cyt aa₃ in this range.⁶ Thus it is possible to separately assess hemodynamic changes and cytochrome reactions or their interactions in response to these changes. In addition, because of the relative transparency of biological tissue to NIR spectrum it is possible to detect photons that have passed through 8 cm or more of tissue thus rendering it feasible to transilluminate the preterm infant's head from one side to another. Although the principle of NIRS involves similar principle used in the pulse oximeter, NIR monitoring provides information on the actual delivery of oxygen to the tissues (more importantly to the brain) rather than in peripheral arterial blood. When the NIR beam passes through tissues, a decrease in signal intensity results from the absorbance of chromophores (HbO₂ and Hb-) and oxidised copper moieties of Cyt aa₃ in its path (Figure 1). Depending on the characteristics of the lasers used in different near-infrared spectroscopes the light spectrum will vary slightly. Generally the light spectrum of special interest is 750-920 nm. At the lower end of this spectrum Hb- has greater absorbance and at the higher end of the spectrum HbO₂. The point of equal light absorbance is 810 nm and is termed the "Isobestic point". The sum of Hb- and HbO₂ indicate tissue blood volume (By). Absorbance band for the copper moieties of cytochrome-c-oxidase (Cyt aa₃), is 780-900 nm with maximum absorbance at 840 nm. Changes in optical absorption are measured in optical densities (arbitrary termed Vander Units by Jobsis) and can be converted into concentration changes (in mmol/L) of these chromophores by employing a modification of the Beer-Lambert principle.^7,8 The NIRS principally provides 4 traces to indicate the tissue (regional) contents of HbO₂, Hb-, B V, and redox state of Cyt aa₃. These traces provide trends viz, they are linearly related to changes in concentration but not absolute concentration of monitored parameters. However, with additional data on simultaneous circulatory status (changes in arterial oxygen saturation (SaO₂) ), tissue blood flow and blood volume can be obtained.^9-11

Fig. 1 Absorption spectra of oxygenated (HbO2) and deoxygenated (Hb) hemoglobin and the oxidised minus reduced difference spectrum of cyto-chrome c oxidase (aa3).15 Reproduced with permission from Elsevier Science Publishers.

The basic system of NIRS consist of four laser diodes and fiberoptic means of monitoring one area of the head, or other organs, using either a transillumination (placed on opposite side of the head as in premature infants or small animals) or reflectance mode (placed close together at acute angles to each other as in term infants or larger body parts such as muscle). The light emanating from the tissue traversed by the photons is captured by a second optode and is either carried by fiberoptics to the photomultiplier in NIRS console or detection takes placed in the detecting optode directly on the skin. The laser diodes are addressed for brief periods in rapid, sequential alternation. After demolution the individual wavelength signals are averaged over a larger fraction of a second. The four biological signals (HbO₂, Hb-, BV, and Cyt aa₃) are calculated by means of algorithms obtained by earlier experimentation in which only one of these parameters was varied at a time.¹² More detailed description of the instrumentation and relevance of signals may be obtained from several publications.^1,13-16

In recent years, attention has been focused on the quantification of NIRS measurements. This requires an estimation of optical pathlength through the tissues. Because of multiple scattering, this pathlength is considerably longer than the distance between the sites of entry and exit. Using the time of flight (TOF) of an ultrashort pulse of infrared light as it traverse the particular tissue of interest (head) Delpy and his colleagues were able to obtain the dimensions of the NIR path which they termed "Pathlength factor" (B). The value for B has been found to vary from an average of 3.9 ± 0.6 by van der Zee, et al¹⁷ to 4.4 ± 0.3 by Delpy, et al.⁸ This factor (B) can then be applied to the modification of the Beer-Lambert equation⁸ and used to calculate the quantification of the NIRS data. Quantification of NIRS data on the neonate's head is possible with the optical fibers positioned at different angles provided the distance of the optodes is greater than 2.5 cm.¹⁸ TOF analysis has also been applied to phase-shift techniques to the delay in moment of occurrence of peak photon flux described by Sevick et al¹⁹ as time-resolved spectroscopy. It is to be noted that measurement of optical pathlength of all infants to be studied is still not feasible, differences between the very low birth weight infants, premature infants and term infants have still to be properly defined. Further pathlength measurements using different optode geometrics should allow quantitative observations to be made in infants of different gestation ages and whether they are small for dates, appropriate or large for gestational age.

The ability of the NIRS to simultaneously provide information on the redox state of Cyt aa₃ with hemodynamic changes in the microcirculation adds an important dimension to the usefulness of the technique. Cyt aa₃ is the terminal member of the mitochondrial respiratory chain. Mitochondrial respiration is the predominant means by which cells utilizes oxygen. Cyt aa₃ catalyses 90% of all O₂ used in the body and more than 95% of the fraction used by the brain leading to the reduction of oxygen to water according to the redox reaction O₂ + 4H⁺ + 4e^- _____ 2H₂O. Transport of reducing equivalents or electrons, derived mainly from the redox reactions in the tricarboxylic cycle (Krebs cycle), through the respiratory chain provides the free energy for conversion of ADP to ATP by the process of oxidative phosphorylation (Figure 2). Thus cytochrome c oxidase plays a key role in the mitochondrial reaction that couples these redox reactions to oxygen utilization to the production of high-energy phosphates (ATP) for cell maintenance, function and growth. Lack of oxygen or the introduction of inhibitors such as cyanide or carbon monoxide would interfere with proper respiratory-chain function and lead rapidly to cellular malfunction and injury.

Fig. 2 Diagramatic representation of the electron transfer of reactions down the mitochondrial respiratory chain. The electron transfer from substrate to oxidation of cytochrome c oxidase provides the free energy for conversion of ADP to ATP.26 Reproduced with permission from Elsevier Science Publishers.

Earlier studies were largely focused on the effect of hypoxia (or oxygen sufficiency) Table. The data obtained from these studies indicate that the Cyt aa₃ optical tracings invariably tracks the HbO₂ tracings; in other words when HbO₂ increased the Cyt aa₃ also became more oxidised and vise versa. Questions arose as to whether the optical tracings for Cyt aa₃ were actually due to Cyt aa₃ or that they represent "cross talk" between the two signals. Studies by Sylvia et al (1982)²⁰, Piandadosi and Jobsis-VanderVliet FF (1984)²¹ in animals (Sprague-Dawley rats) replacing blood with fluorocarbon (Fluosil 43) showed that the Cyt aa₃ optical tracings were independent of HbO₂. Additional studies by Sylvia AL et al. (1985)²² of rats subjected to graded arterial hypoxia demonstrated that the NIRS signals of Cyt aa₃ correlated with cerebral concentrations of pyruvate, lactate, phosphocreatine, ATP and ADP changes. Direct inhibition of Cyt aa₃ by cyanide in bloodless rats was reported by Piantadosi CA and Sylvia AL.²³ More recent studies by Wu et al,²⁴ demonstrated that the entrance of bilirubin into neuronal cells is followed by increased reduction of Cyt aa₃ independent of the Hb signals. Other studies by these investigators,²⁵ using an animal model (rats) demonstrated that Cyt aa₃ was also markedly reduced by potassium cyanide (KCN) and was associated with decreased glutathione in the brain independent of Hb signals. These studies validate the NIRS optically derived Cyt aa₃ signals.

The ability of the NIRS to provide noninvasively, continuous, real-time information on cerebral hemodynamics and tissue oxygenation makes this a potentially useful instrument to monitor hypoxia and its effects on cerebral metabolism. Thus early studies with the NIRS were focused on characterization of oxygen insufficiency. In fact the Niros-scope made by Josis stands for near infrared oxygen sufficiency spectroscope. Thus the earliest applications of the NIRS in neonatal medicine by Brazy and her colleagues^3-5 were to monitor pathophysiologic responses in cerebral HbO₂, Hb-, BV and mitochondrial Cyt aa₃ in infants who were at risk for hypoxia, due to diseased states (e.g. respiratory distress) or to handling or various procedures involved in the care of the infant. The findings are in agreement with our studies (using newborn piglets) that demonstrated that hypoxia caused reduction in HbO₂, increased Hb-, and BV with concomitant increased in reduction of Cyt aa₃²⁶ (Figure 3). Additionally with graded degrees of hypoxia, these changes became more marked (Figure 4). Studies by Sylvia, et al²¹ showed that the redox state of Cyt aa₃ at different F_iO₂ was directly related to in vitro measured changes in cortical metabolites known to reflect energy production. Thus the redox state of cerebral Cyt aa₃ would therefore reflect the state of cerebral energy metabolism.

Fig. 3 Representative optically derived tracings showing the effect of transition from normoxia (room air) to hyperoxia (100% O2) to hypoxia (100 N2) and back to hyperoxia. Note Cyt aa3, tHbO2 and tBV changes tended to overshoot with change from hypoxia to hyperoxia.26 Reproduced with permission from Elsevier Science Publishers.

Fig. 4 Depicts the changes in tissue HbO2, Hb-, BV, and Cyt aa3 with graded changes in FiO2. Reproduced with permission from Elsevier Science Publishers.

Identification of the redox state of Cyt aa₃ had been shown to be of particular usefulness in identifying, so called undedected (silent), episodes of cellular hypoxia as found in association with a large patent ductus arteriosus (PDA) during conditions of reduced cardiac output.^4-5 These studies also showed that recovery of Cyt aa₃ to baseline may not parallel the recovery of Hb oxygenation.⁶ The implications of this delay will need to be elucidated. It is quite possible that this delay in recovery may signal neuronal dysfunction or impending cellular injury. The effects of indomethacin used in the closure of the PDA were studied by several investigators.^27-29 These investigators found that following indomethacin administration, cerebral blood flow, oxygen delivery, blood volume, and reactivity of blood volume to changes in arterial carbon dioxide tension fell sharply, with concomitant increased reduction of Cyt aa₃. These changes in cerebral perfusion may therefore result in cerebral haemorrhage or cellular damage.

Another potential use of the NIRS had been applied to monitoring RDS during surfactant replacement therapy. Cerebral HbO₂ fell transiently, while Hb- and BV increased during the drug administration and then recovered quickly.^30-32 The significance of these fluctuations in hemodynamics is not quite clear at present. Studies by Greisen³³ and his associates found that these changes were associated with marked depression in EEG activity lasting for 10 - 20 mm after surfactant administration. Whether these findings have a permanent effect are unknown.

The effects of positive and negative pressure ventilation on cerebral hemodynamics were recently studied in infants with RDS with the use of NIRS.³⁴ Continuous negative extrathoracic pressure caused a median decrease in cerebral volume of 0.14 ml/100 ml brain compared with no respiratory support. HbO₂ and Hb- both decreased, suggesting increased venous drainage as the main effect. Intermittent positive pressure ventilation (IPPV) also caused a median reduction in cerebral BV of 0.06 ml/100 ml brain. Hb- was found to increase while HbO₂ decreased. The increase in Hb- suggests decreased venous drainage due to increase in intrathoracic pressure.³⁵ This may lead to increase in venous blood within the cerebral circulation. HbO₂ changes may be associated with fluctuations in CBV during the IPPV circle.³⁶ Further, deLemos and Tomasovic³⁷ showed transmission of positive airway pressure to the thorax increased as lung compliance improved with resolving RDS. Applications of the NIRS to monitor cerebral blood flow with various forms of ventilatory settings and ventilators for infants of differing sizes and diseases would be useful to arrive at parameters that would result in maximal effectiveness but decreased risks for cerebral hemorrhage.

The possibility of applying the NIRS to obtain insights into conditions apart from hypoxia (or oxygen sufficiency) has recently been reported by Wu and his colleagues in several studies aimed at characterizing the pathophysiological changes in cerebral hemodynamics and mitochondrial metabolism, as reflected by the redox state of Cyt aa₃, in pathogenic Escherichia coli (E. coli) sepsis.^38-40 Their studies found that following infusion of E. coli into the blood stream there was an initial hyperdynamic reaction with increases in cerebral BV, Hb- and Cyt aa₃ which lasted from 1-2 hours. Subsequently, the BV and HbO₂ decreased markedly while Hb- increased and Cyt aa₃ became more reduced. These data suggest that the initial hyperdynamic reaction in cerebral microcirculation with increase in oxygen uptake leading to increased oxidation of Cyt aa₃ (thereby cell metabolism). As sepsis continues the decrease in cerebral circulation lead to inadequate oxygen for the increased metabolism of brain cells resulting ultimately in neuronal injury and cell death. These studies also showed that pretreatment with dexamethasone as well as monoclonal antibody (specific anti 018) can modulate or prevent the changes in cerebral microcirculation and maintain the steady redox state of Cyt aa₃.

Observations with the NIRS have recently been extended to the fetus during the intrapartum period.⁴¹ Special optical probes, designed by Peebles and his colleagues, that can be pass through the cervix and applied to the fetal scalp to provide continuous information on the cerebral hemodynamic changes during uterine contractions.^42,43 Assessment of the usefulness of interventions during labor (e.g. oxygen or oxytocin) may also be obtained.^44,45

Thus data from animal and clinical studies in research settings indicate that the near infrared spectroscopy has great potential as a noninvasive device for providing continuous, real-time information on cerebral hemodynamics, oxygenation, and tissue oxygen sufficiency. The technique can be applied to characterize the evolution of pathophysiologic responses during diseased states and the effectiveness of treatment modalities. Predictive value of the results obtained from such monitoring needs to be assessed. Advances in development for the quantification of the measured parameters will still need to be refined.

Acknowledgements

The author gratefully acknowledge the invaluable guidance and assistance by Drs. Frans F Jobsis, Avis L Sylvia, Kwan S Kim, Manual Durand, Won S Park, Rowena G Cayabya and Maria-Ellen S Mendoza in various studies cited in this paper.

References

1. Jobsis FE. Non-invasive, infra-red monitoring of cerebral and myocardial oxygen sufficiency and circulatory parameters. Science 1977;198:1264-7.

2. Jobsis FF, Keizer JH, LaManna JC, Rosenthal M. Reflectance spectrophotometry of cytochrome aa₃ in vivo. J Appl Physiol 1977;43:858-72.

3. Brazy JE. Effects of crying on cerebral blood volume and cytochrome aa₃. J Pediatr 1988;112:457-61.

4. Brazy JE, Lewis DV. Changes in cerebral blood volume and cytochrome aa₃ during hypertensive peaks in preterm infants. J Pediatr 1986;108:983-7.

5. Brazy JE. Near-infrared spectroscopy. Clin Perinatol 1991;18:519-34.

6. Brazy JE, Lewis DV, Mitnick MH, Jobsis FE. Noninvasive monitoring of cerebral oxygenation in preterm infants: Preliminary observations. Pediatrics 1985;75:217-25.

7. Wray S, Cope M, Delpy DT, Wyatt JS, Reynolds EOR. Characterization of near infrared absorption spectra of cytochrome aa₃ and haemoglobin for noninvasive monitoring of cerebral oxygenation. Biochim Biophys Acta 1988;33:184-92.

8. Delpy DT, Cope M, van der Zee P, Arridge S, Wray S, Wyatt JS. Estimation of optical pathlength through tissue from direct time of flight measurement. Phys Med Biol 1988;33:1433-42.

9. Edwards AD, Wyatt JS, Richardson EC, Delpy DT, Cope M, Reynolds EOR. Cotside measurement of cerebral blood flow in ill newborn infants by near infrared spectroscopy. Lancet 1988;ii:701-2.

10. Wyatt JS, Cope M, Delpy DT, et al. Quantitation of cerebral blood volume in newborn human infants by near infrared spectroscopy. J Appl Physiol 1990;68:1086-91.

11. Wyatt JS. Near infrared spectroscopy : Investigation and assessment of perinatal brain injury. Biol Neonate 1992;62:290-4.

12. Jobsis-Vandervliet FF. Niros-scopy: Non invasive near infrared monitoring of cellular oxygen sufficiency in vivo. Adv Exp Med Biol 1986;191:833-42.

13. Brazy J. Cerebral oxygen monitoring with near infrared spectroscopy: Clinical application to neonate. J Clin Monit 1991;7:325-34.

14. Jobsis-VanderVliet FF, Piantadosi CA, Sylvia AL, Lucas SK, Keizer HH. Near-infrared monitoring of cerebral oxygen sufficiency. 1. Spectra of cytochrome c oxidase. Neuro Res 1988;10:7-18.

15. Jobsis-VanderVliet FF. Near infrared monitoring of cerebral cytochrome c oxidase : Past and present (and future ?) in Fetal and Neonatal PhysiologicaL Measurements, Lafeber HN (ed) Elsevier Science Publishers BV. 1991;52.

16. Wray S, Cope M, Delpy DT, Wyatt JS, Reynolds EOR. Characterization of the near infrared absorption spectra of cytochrome aa₃ and hemoglobin for non-invasive monitoring of cerebral oxygenation. Bioch et Biophys Acta 1988;933:184-92.

17.van der Zee P, Cope M, Arridge SR, et al. Experimentally measured optical pathlength for adult head, calf and forearm, and the head of newborn infant as a function of inter optode spacing. Adv Exp Med Biol 1991;316:143-53.

18.Wyatt JS. Near-Infrared Spectroscopy in Asphyxial Brain Injury. Cl Perinatol 1993;20:369-78.

19. Sevick EM, Chance B, Leigh J, Nioka S, Mans M. Quantitation of time- and frequency-resolved optical spectra for the determination of tissue oxygenation. Anal Biochem 1991;195:350-1.

20. Sylvia AL, Proctor HJ, Goldsmith MM, Jobsis FE. Exchange transfusion with Fluosol 43: In vivo assessment of cerebral cytochrome coxidase redox state. J Trauma 1982;22:815-9.

21. Piandadosi CA, Jobsis-VanderVliet FE. Spectrophotometry of cerebral cytochrome aa₃ in bloodless rats. Brain Res 1984;305:89-94.

22. Sylvia AL, Piantadosi CA, Jobsis-VanderVliet FF. Energy metabolism and in vivo cytochrome c oxidase redox relationships in hypoxic rat brain. Neurol Res 1985;7:81-8.

23. Piandodosi CA, Sylvia AL. Cerebral cytochrome aa₃ inhibition by cyanide in bloodless rats. Toxicology 1984;33:67-79.

24. Wu PYK, Jobsis FF, Sylvia AL. Cerebral changes in cytochrome aa₃ as an indicator of bilirubin uncoupling of oxidative phosphorylation. Pediatr Res 1991;29:240A.

25. Wu PYK, Kannan R, Cayabyab RG, Mendoza MES. Effects of cyanide on brain mitochondrial cytochrome oxidase. Pediatr Res 1994;4:389A.

26. Wu PYK, Jobsis-VanderVliet FF. In vivo brain cytochrome c oxidase redox responses to changes in cerebral oxygenation. In Fetal and Neonatal Physiological Measurements, Lafeber HN ed. Elsevier Science Publishers B.V. 1991;73-6.

27. Edwards AD, Wyatt JS, Richardson C, et al. Effects of indomethacin on cerebral hemodynamics in very preterm infants. Lancet 1990;335:1491-5.

28. McCormick DC, Edwards D, Brown GC, et al. Effect of indomethacin on cerebral oxidized cytochrome oxidase in preterm infants. Pediatr Res 1993;33:603-8.

29. Benders MJNL, Dorrepaal CA, van de Bor M, van Bel E. Acute effects of indomethacin on cerebral hemodynamics and oxygenation. Biol Neonate 1995;68:91-9.

30. Fahnenstich H, Schmidt S, Spaniol S, Kpwalcski S. Relative changes in oxyhemoglobin, deoxyhemoglobin and intracranial blood volume during surfactant replacement therapy in infants with respiratory distress syndrome. Dev Pharmacol Ther 1991;17:150-3.

31. Edwards AD, McCormick DC, Roth SC, et al. Cerebral hemodynamic Effects of treatment with modified natural surfactant investigated by near infrared spectroscopy. Pediatr Res 1992;5:532-6.

32. Dorrepaal CA, Benders MJNL, Steeendijk P, van del Bor, van Bel F. Cerebral hemodynamics and oxygenation in preterm infants after low-vs. high-dose surfactant replacement therapy. Biol Neonatl 1993;64:193-200.

33. Skov L, Bell A, Greisen G. Surfactant administration and cerebral circulation. Biol Neonate 1992;61(suppl 1):3 1-6.

34. Palmer KS, Spencer SA, Wickramasinghe YABD, Wright T, Southall DP, Rolfe P. Effects of positive and negative pressure ventilation on cerebral blood volume of newborn infants. Acta Padiatr 1955:84:132-9.

35. Robotham J, Scharf. Effects of positive and negative pressure ventilation on cardiac performance. Clin Chest Med 1983;4:161-7.

36. Leahy FAN, Durand M, Cates D, Chernick V. Cranial blood volume changes during mechanical ventilation and spontaneous breathing in newborn infants. J Pediatr 1982;101:984-7.

37. deLemos RA, Tomasosvi JJ. Effects of positive pressure ventilation on cerebral blood flow in newborn infant. Clin Perinatol 1978;5:393-409.

38. Mendoza MES, Cayabya RG, Park WS, Wu PYK, Kim KS. Responses of cerebral microcirculation and mitochondrial cytochrome oxidase redox state to E coli. Pediatr Res 1994;35:383A.

39. Cayabya RG, Mendoza MES, Wu PYK, Kim KS. Effects of dexamethasone on responses of cerebral microcirculation and mitochondrial cytochrome oxidase redox state to E coli. Pediatr Res 1994;35:1755A.

40. Cayayab RG, Mendoza MES, Wu PYK, Kim KS. Role of monoclonal antibody of cerebral microcirculation and mitochondrial cytochrome oxidase redox state in E coli sepsis. Pediatr Res 1994;35:1756A.

41. Wyatt JS, Peebles DM. Near infrared spectroscopy and intrapartum fetal surveillance. In: Spencer JAD, ed. Intrapartum fetal surveillance. London; Royal College of Obstetricians and Gynaecologists, 1993:329-46.

42. Peebles DM, Edwards AD, Wyatt JS, et al. Changes in human fetal cerebral hemoglobin concentration and oxygenation during labor measured by near infrared spectroscopy. Am J Obstet Gynecol 1992;166:1369-73.

43. Peebles DM, Spencer JAD, Edwards AD, et al. Relation between frequency of uterine contractions and human fetal cerebral oxygen saturation studied during labor by near infrared spectroscopy. Br J Obstet Gynecol 1994;101:44-8.

44. Aldrich CJ, Wyatt JS, Spencer JAD, Reynolds EOR, Delpy DT. The effect of maternal oxygen administration on human fetal cerebral oxygenation measured during labor by near infrared spectroscopy. Br J Obstet Gynaecol 1994;101:509-13.

45. Peebles DM, Edwards AD, Wyatt JS. Effect of oxytocin on fetal brain oxygenation during labor. Lancet 1991;338:254-5.