Human Germinal Matrix: Venous Origin of Hemorrhage and Vascular Characteristics Hani S. Ghazi-Birry, William R. Brown, Dixon M. Moody, Venkata R. Challa, Steven M. Block, and David M. Reboussin PURPOSE: To examine the vascular supply and architecture of the germinal matrix in the preterm neonatal brain and to determine whether veins or arterioles are the source of germinal matrix hemorrhage. METHODS: Brains from eight preterm neonates (24 to 35 weeks’ gestation) and two full-term infants were fixed in alcohol, embedded in celloidin, sectioned at 100- and 500-mm thicknesses, stained for alkaline phosphatase, and examined with light microscopy. High-resolu- tion contact radiographs of 500-mm-thick sections were also mounted on glass slides for micro- scopic examination. RESULTS: The upper and middle regions of the germinal matrix are supplied by branches of the lateral striate arteries, whereas the inferior part is supplied by branches of the recurrent artery of Heubner. In brain sections from four of the preterm infants, we found 15 circumscribed hemorrhagic foci within the germinal matrix. The largest was 5 mm in diameter; the smallest, 1 mm. All hemorrhages but one were closely associated with veins, with significant involvement of the perivenous space. The other hemorrhage appeared to be associated with an arteriole. In term and preterm infants, we found no arteriolar-to-arteriolar shunts, precapillary arteriolar-to-venules shunts, or vascular rete. At all gestational ages, the terminal vascular bed had only conventional branchings and connections. CONCLUSION: In preterm neonates, staining for endogenous alkaline phosphatase allows visual differentiation between afferent and efferent ves- sels. Germinal matrix hemorrhage in preterm neonates is primarily venous in origin. A hemorrhage can tunnel along the venous perivascular space, collapsing the vein and rupturing the tethered connecting tributaries. Extravasation of blood from the arterial circulation appears to be much less common. Index terms: Cerebral hemorrhage; Infants, newborn; Pathology AJNR Am J Neuroradiol 18:219–229, February 1997 Hemorrhages involving the neonatal brain are show a considerable decline in the proportion of among the most frequent and serious events such lesions and a corresponding marked in- affecting the newborn. Although subdural he- crease in the frequency of germinal matrix hem- matoma was formerly the most common type of orrhage (1). The primary reasons for this intracranial hemorrhage, probably as a result of change are the marked reduction in the number adverse obstetric conditions, more recent data of subdural hematomas resulting from im- proved obstetric management of the second stage of labor and a relative increase in cases of Received May 20, 1996; accepted after revision August 19. germinal matrix hemorrhage associated with Supported by NIH NS20618 Jacob K. Javits Neuroscience Investigator Award (D.M.M.) and the March of Dimes Birth Defects Foundation Grant the increased survival of premature infants with #6-FY95–223 (W.R.B.). very low birth weight, who are supported by From the Department of Radiology (H.S.G-B., W.R.B., D.M.M.), the advanced neonatal intensive care. Currently, Program in Neuroscience (W.R.B., D.M.M.), the Department of Pathology germinal matrix hemorrhage is the most fre- (W.R.B., V.R.C.), the Department of Pediatrics (S.M.B.), and the Depart- quent and important brain abnormality in the ment of Public Health Sciences (D.M.R.), Bowman Gray School of Medicine of Wake Forest University, Winston-Salem, NC. neonatal period. Address reprint requests to Dixon M. Moody, MD, Department of Radi- Germinal matrix hemorrhage occurs primar- ology, Bowman Gray School of Medicine, Medical Center Blvd, Winston- ily but not exclusively in premature neonates of Salem, NC 27157. very low birth weight. The recent large multi- AJNR 18:219–229, Feb 1997 0195-6108/97/1802–0219 center study coordinated by the National Insti- © American Society of Neuroradiology tute of Child Health and Human Development 219 220 GHAZI-BIRRY AJNR: 18, February 1997 reported a 45% overall occurrence rate for ger- variously with such noncommittal terms as ves- minal matrix hemorrhage in preterm low-birth- sels, sinusoids, or channels. weight neonates (2). Since premature infants One method for differentiating venous from made up 9.7% of live-born infants in the United arterial vessels is alkaline phosphatase histo- States in 1988, approximately 400 000 prema- chemical staining. This research procedure, for- ture births would have occurred in 1995, and merly used for the study of normal adult angio- 40 000 of such infants would weigh less than architecture, has been used to investigate adult 1500 g at birth; neonates with germinal matrix vascular-based central nervous system abnor- hemorrhage therefore constitute an enormous malities and to study brain vascular morphol- medical, social, and financial challenge for the ogy in preterm neonates of very low birth weight nation (3). Additionally, improved obstetric and (12). In the present study we used alkaline neonatal techniques have led to the survival of a phosphatase histochemical staining and other not-insignificant percentage of extremely pre- techniques to identify various characteristics of mature neonates. Indeed, reports describing the vascular supply and architecture of the ger- survival of premature neonates with gestational minal matrix in the brain of these preterm, low- ages of 22 to 24 weeks are not infrequent, and birth-weight infants. Our purpose was to deter- such infants have the highest risk for germinal mine whether the large germinal matrix matrix hemorrhage. Moreover, a significant per- channels are venules, arterioles, or capillaries centage of preterm neonates with germinal ma- and to ascertain which types of vessels are the trix hemorrhage eventually contract cerebral source of the germinal matrix hemorrhage. palsy, which is a consequence of germinal ma- trix hemorrhage, its complications, or the asso- Materials and Methods ciated lesions of periventricular leukomalacia or periventricular hemorrhagic infarction (4). Materials Therefore, the significance of this condition We studied brains obtained at autopsy from eight pre- cannot be overemphasized. term neonates of very low birth weight (postconception To reduce the prevalence of germinal matrix ages, 24 to 35.5 weeks) and from two full-term infants. hemorrhage, one must understand its patho- Four of the preterm neonates had clinical diagnoses of genesis, and to do so requires detailed knowl- germinal matrix hemorrhage, which were subsequently edge of the anatomy of the vascular system confirmed with pathologic examination of the brain. Typ- within the germinal matrix in the preterm infant ically, various combinations of the following prenatal risk as well as knowledge of the vascular etiologic factors and postnatal clinical abnormalities were present source of germinal matrix hemorrhage. At and may have contributed to the demise of the patients: present, some speculate that the vascular maternal drug abuse, maternal insulin-dependent diabetes source of germinal matrix hemorrhage in the mellitus, antepartum hemorrhage, uteroplacental insuffi- ciency, respiratory distress syndrome, hyaline membrane premature infant of very low birth weight is ve- disease, pneumothoraces, severe hypoxia, hypercapnia, nous (5–8), whereas others believe it is capil- acidosis, severe hypotension, prolonged bradycardia, sep- lary or arterial (9–11). Precise description of sis, anemia, thrombocytopenia, and other complications neonatal brain vasculature, generally, and of of prematurity. Both full-term infants died as a conse- the bleeding source, specifically, has remained quence of congenital heart malformations and associated elusive because of the difficulty in classifying complications. Details of gestational and postconception the immature vascular structures in the germi- ages at death, birth weight, brain weight, Apgar scores, nal matrix. In the deep brain microvasculature and germinal matrix hemorrhage grade for each patient of the premature infant, it is difficult to identify are presented in the Table. supporting wall tissue (collagen in veins, smooth muscle in afferent vessels). The cause Methods of germinal matrix hemorrhage cannot be es- tablished definitively without first classifying Immediately after autopsy, the neonatal brains were placed in cold 70% alcohol for at least 10 days for fixation. vessels into functional categories (ie, arterioles, Formalin fixation was avoided because it destroys the capillaries, or veins) histologically. Although endogenous alkaline phosphatase used by our staining certain large, immature vessels in the germinal technique. Subsequently, a whole-brain slice approxi- matrix seem to be implicated in germinal matrix mately 1 cm thick, incorporating both hemispheres, was hemorrhage, their morphologic status remains obtained from each brain at and posterior to the foramen of obscure, and investigators have classified them Monro and included the thickest mass of germinal matrix AJNR: 18, February 1997 GERMINAL MATRIX 221 Demographic data and germinal matrix hemorrhage (GMH) status Gestational/ Birth Apgar Brain GMH GMH Patient Postconception* Weight, g Score† Weight, g Present Grade‡ Age, wk 1 550 24/24 2/5 76 Yes II 2 660 24/24 1/2 52 No ... 3 635 24/33 2/7 180 Yes II 4 760 26/26 1/6 96 Yes III 5 454 28/28 5/6 88 No ... 6 1310 29/29 ... 185 No ... 7 1480 31.5/32.5 1/3 220 Yes II 8 3250 35/35.5 2/2 280 No ... 9 2870 38/38.5 8/8 330 No ... 10 3300 40/43 9/9 420 No ... * Postconception age at death. † Apgar scores are at 1 and 5 minutes; patient 6 was born at home, so no score was available. ‡ GMH grade is according to the criteria of Papile et al (20). as well as basal ganglia, thalamus, centrum semiovale, acetate/light green, Gill’s hematoxylin). Our technique of and cortex. The brain slices were then dehydrated in as- differential visualization of afferent vessels from veins has cending grades of ethanol, embedded in celloidin, and been tested in different species and different organs and it serially sectioned at 100 mm, and 500 mm on a base sledge has proved to be a reliable and robust procedure.