MICROSCOPY RESEARCH AND TECHNIQUE 45:339340 (1999)

Introduction to Histochemical Studies on Human Fetal Brain Development

NORBERT ULFIG
Neuroembryonic Research Laboratory, Department of Anatomy, University of Rostock, D-18055 Rostock, Germany

During brain ontogenesis, the immense diversity of the neural and glial phenotype emerges and neural circuitries are established. Understanding the events of brain development is a prerequisite to interpret developmental disorders. The histological structure of the immature brain differs substantially from that of the mature and it continuously changes during maturation. In order to determine the causes of abnormalities in the developmental program, exhaustive data on normal development are required. The bulk of information that is so far available on brain development, however, is derived from animal studies. It was, therefore, a challenge to collect review as well as original articles dealing with various aspects of human brain development. Several major developmental events are considered in the articles of this special issue. The rst major step in brain ontogenesis critical in shaping the brain is the generation of neurons and most glial cells in the neuroepithelium lining the ventricles. The number of cells generated in a particular portion of the neuroepithelium depends on the number of precursor cells, the duration of cell cycles, and the number of cell divisions. Additionally, apoptosis in the neuroepithelium may also determine the number of cells produced. Both processes, proliferation and cell death, have been investigated in human brains of the second trimester; their link to each other and their possible relationship to subsequent developmental steps are discussed by Simonati et al. In their investigation, they also included an extraventricular proliferation site: In the cerebellum, cells from the neuroepithelium migrate as precursors to form a secondary proliferative zone. The latter is characterized by a late proliferation period that extends into postnatal life. Recently, the processes of proliferation have been shown not to be restricted to the pre- or perinatal period. Neurons are generated throughout adulthood within the hippocampal formation. The paper by Del Bigio, providing evidence that hippocampal neurons are newly generated in the human adult, is an important contribution with regard to plasticity in the adult brain. The functional signicance of this neuronal production in the normal adult brain has yet to be elucidated. Components of the CNS not derived from the neuroepithelium are also considered here; i.e., the anterior pituitary gland (see below) and microglia. The enigmatic microglial cells, representing a signicant element of CNS tissue, are generally believed to form from hemopoietic cells. The colonization of the brain by microglial stem cells, phenotypic appearance, and functional roles of microglia and their interaction with neuroepithelial cells are, among others, discussed in the extensive review by Rezaie and Male.
1999 WILEY-LISS, INC.

Young postmitotic neurons have to migrate from the proliferative zones towards their denitive positions. Migration is guided by long radially oriented glial bers that provide scaffolding for migrating neurons. The latter follow the radial pathways via surface-mediated interactions. Itoh et al. describe the expression of a novel peroxisomal enzyme (D-bifunctional protein) in neurons during early fetal stages. As a deciency of peroxisomal enzymes has been reported to lead to migration disorders, it is plausible to assume that the D-bifunctional protein plays a signicant role in the process of migration. Mainly following cell migration, growth and differentiation of neurons occur. These processes underlying the assembly of mature brain structures comprise cell transformation and cell interactions. Undifferentiated neurons develop into polarized nerve cells with a complex shape that is determined by characteristics of the soma, the dendritic tree, and the course of the axon. During differentiation, the cellular apparatus for neuronal function is formed and the synthesis of neurotransmitters, other neuroactive substances, and receptor proteins is initiated. The programmed neuronal phenotype can be altered by various factors of the microenvironment. The latter can also determine alignment and orientation of neurons. To assess neuronal differentiation in the human, it is desirable to get a survey on the spatial and temporal distribution of neurotransmitters and neuropeptides. In their review, Chan and Yew discuss morphological data on the distribution patterns of neuroactive substances and their early appearance with functional aspects, such as the establishment of neuronal circuitries, the roles of these substances in process outgrowth and synapse formation, their signicance as neurotrophic factors, and interactions between the various substances. Nitric oxide, a novel messenger molecule, is expressed early during development. Judas et al., therefore, summarize various developmental roles of nitric oxide, for instance in synaptogenesis or cell death, based on morphological data and the area-specic transient expression of nitric oxide. Transformation of transient patterns is likely to indicate developmental changes of circuitry elements. Certain events during neuronal differentiation, such as process outgrowth, are related to intracellular calcium concentrations. Such events may, therefore, be mediated by calium-binding proteins, such as calbindin and calretinin. Setzer and Ulg describe that these two proteins are expressed in distinct neuronal subpopulations of the fetal amygdala and, thus, point out that certain nerve cell classes can be subdivided. The neuro-

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chemical heterogeneity may be associated with differential connections of neuronal subclasses. The formation of neuronal connections may be regulated by proteins that have so far been investigated only infrequently. Arii et al. describe the distribution pattern of the glycoprotein telencephalin, a cell adhesion molecule that is solely expressed in the telencephalon. The authors postulate an involvement of telencephalin in the late phase of synaptogenesis and dendritic maturation. Afferents are likely to exert a regulatory inuence on differentiating neurons. Thus, neurotransmitters interact via their respective receptors to inuence neuronal survival and differentiation. Neuronal populations are reached by various afferent systems with different neurotransmitters occurring in spatio-temporal orders. Interactive effects must, therefore, be taken into account. Early in development, a widespread and diffuse innervation of the cerebral cortex is prominent. The early ontogenesis of monoaminergic neurons and the precocious development of their projections are thoroughly reviewed by Verney. She interestingly points out the specic charcteristics unique to humans and transient features reecting reorganizational events. Synapses can reliably be demonstrated by electron microscopy, which, however, depends upon very short postmortem delays. The establishment of time-tables of synaptogenesis is rendered possible with the aid of mapping the expression of synapse-related molecules. Postsynaptic densities contain the protein AKAP79, which is detectable in an area-specic manner in the human fetal amygdala. Ulg and Setzer nd that AKAP79 is expressed in various neuronal types and conclude that it cannot be assigned to denite amygdaloid circuitries. Such results can also provide the basis for investigations on the elimination of synapses as a process of remodelling of connections. The last two papers of this issue mainly deal with disturbances of ontogenetic key events. So far it is not known when the transitional bipotential cell of neuroepithelium becomes irreversibly committed to neuronal or glial differentiation. Blu mcke et al. postulated the existence of a bipotential precursor cell that is the starting point of glio-neuronal tumors. The latter are regarded to be a morphological substrate of chronic temporal lobe epilepsy in young patients. An overview on the development of the anterior pituitary gland, which is derived from the roof of the pharynx, is given by Hori et al. They particularly focus on disturbances of histogenesis and cell migration (ectopia) with special reference to tumorigenesis. The data presented here obtained from histochemical studies on the human fetal brain establish baselines for the evaluation of pathological material in more detail. So far, the morphological substrate of a number of developmental disturbances has been investigated only fragmentarily most probably due to the fact that changes are often rather subtle. Such alterations can only be detected using specic histochemical techniques that are so far not routinely applied. Their application in neuropathology, however, requires a detailed knowledge on the normal structure. Undoubtedly, this special issue has made a contribution to this approach, which would, moreover, provide insight in the pathogenesis of developmental disorders. It is a formidable task to further apply concepts in developmental neurobiology to an understanding of the origins of disorders as advances in the prevention of disturbances are based on a thorough and complete understanding of the alterations occurring in a disease. ACKNOWLEDGMENTS The Guest Editor thanks Dr. John E. Johnson, Jr., Editor-in-Chief of Microscopy Research and Technique, for enabling this special issue, all the authors for their contributions, and Frank Neudo rfer for his valuable assistance in preparing this issue.