Synaptogenesis in the Cerebellum of Offspring Born to Diabetic Mothers

Abstract

There is increasing evidence that maternal diabetes mellitus during the pregnancy is associated with a higher risk of neurodevelopmental and neurofunctional anomalies including motor dysfunctions, learning deficits, and behavioral problems in offspring. The cerebellum is a part of the brain that has long been recognized as a center of movement balance and motor coordination. Moreover, recent studies in humans and animals have also implicated the cerebellum in cognitive processing, sensory discrimination, attention, and learning and memory. Synaptogenesis is one of the most crucial events during the development of the central nervous system. Synaptophysin (SYP) is an integral membrane protein of synaptic vesicles and is considered to be a marker for synaptic density and synaptogenesis. Here, we review the manuscripts focusing on the negative impacts of maternal diabetes in pregnancy on the expression or localization of SYP in the developing cerebellar cortex. We believe that the alteration in synaptogenesis or synapse density may be part of the cascade of events through which diabetes in pregnant women affects the newborn's cerebellum.

INTRODUCTION

Diabetes mellitus during pregnancy period is one of the most common metabolic complications that can be classified into two categories: pregestational or gestational diabetes.[1,2,3,4,5,6,7] This metabolic disorder occurs in 3%–5% of pregnancies.[8,9] Nevertheless, the prevalence of diabetes in pregnancy has been reported to range between <1% to about 14% in different societies.[2,10,11,12] An increasing number of evidence clearly showed that children born to diabetic mothers are in increased risk of fetal and neonatal anomalies including central nervous system (CNS) abnormalities[9,13] which results in increased infant mortality and morbidity rates.[2,7,9,14,15] Previous investigations also report that maternal diabetes is associated with a higher risk of long-lasting neurological impairment that is manifested as deficits in balance and motor coordination, hyperactivity, impairments in attention and memory, and altered social behavior in offspring.[13,16,17,18,19,20,21]

The functional and structural effects of diabetes in pregnancy on the developing CNS have been studied in both human and experimental models.[22,23] Although, the precise mechanisms that diabetes in pregnancy can affect the development and function of the nervous system remain to be defined.[2,5,7,22,23,24] In humans, children from mothers with diabetes during the pregnancy may exhibit abnormalities, which include learning defects, motor difficulties, attention deficit, and also the risk of developing schizophrenia.[19,25,26] Babiker indicated a higher prevalence of developmental delay and behavioral problems in children born to diabetic women. In that study, growth motor skills and language delay were the major development areas of concern they could link between maternal diabetes and development.[17] There is also evidence demonstrating a negative relationship between the performances of the children born to diabetic mothers with the severity of maternal hyperglycemia.[7,27,28] Recent studies found negative effects of maternal diabetes during pregnancy on the development of cerebellar cortex and also the expression and localization of insulin-like growth factor-1 receptor (IGF-1R) and insulin receptor (InsR), as two regulators of CNS development, in the cerebellum and hippocampus of neonatal rats as a probable mechanism for the neurodevelopmental and neurobehavioral impairments observed in diabetic mothers’ offspring.[14,16,29,30,31] Other investigation by Hami et al. also evaluated the effects of maternal diabetes on the developing cerebellum of rats. They estimated cerebellar volume, the thickness and the number of cells in the different layers of the cerebellar cortex. Their results have been shown a significant reduction in the cerebellar volume and the thickness of various developing cerebellar cortex such as external granule layer (EGL), molecular layer (ML), and internal granule layer (IGL) in the offspring born to diabetic animals. They also reported that diabetes in pregnancy disrupts the morphogenesis of developing cerebellar cortex and believed that this dysmorphogenesis may be part of the cascade of events through which maternal diabetes affects motor coordination and social behaviors in offspring.[32] A study by Khaksar et al. also indicated the detrimental effects of diabetes during pregnancy period on the thickness of cerebellar cortex in neonatal rats.[33] Overall, there are multiple lines of evidence suggesting that diabetes in pregnancy can cause neurofunctional and neurostructural abnormalities in the offspring by alteration of many developmental events such as neurogenesis, migration, and differentiation.[16,34]

With regards to the importance of synaptogenesis in developing cerebellum, here, we have reviewed the related articles focusing on the effects of maternal diabetes during pregnancy on the expression or localization of synaptophysin (SYP) in the developing cerebellar cortex of neonatal rats. We believe that the neurodevelopmental and neurofunctional impairments observed in the children born to diabetic mothers may be mediated, at least in part, via alterations in expression and localization of SYP in the developing cerebellum.

DISCUSSION

Synapse and synaptogenesis

Synapses are specialized connections between neurons permitting the controlled transfer of chemical/electrical signals between presynaptic neuronal cells and postsynaptic target neurons.[35] Adequate synapse function is an essential prerequisite of all neuronal processing.[35] True connection between neurons is fundamental to the physiological function of the nervous system,[36,37] and perception, learning, and memory are only possible when the nervous system is functioning normally.[38]

During the development of CNS, synaptogenesis-formation and maturation of synaptic contacts-is one of the most crucial events that is represents the final step of neuronal differentiation.[2,23,39,40,41] Synaptogenesis begins early in the embryo and extends well into postnatal life.[42,43] In mammals, the period and length of newly synapse formation is vary widely from species to species which is in human neocortex occurs during the third trimester of gestation and the first 2 postnatal years, whereas in rodents, the first 2 weeks after birth represent the most active phase of synaptogenesis.[39,40,44,45] Moreover, the formation of a functional synapse is a complex process that involves multiple stages such as axon guidance, synapse formation, synapse maintenance (stabilization), and activity-dependent synapse elimination.[42,46,47,48]

Chemical synapses are best characterized as asymmetric neuronal junctions which are composed of three compartments: The presynaptic bouton, the synaptic cleft and the postsynaptic apparatus.[23,49,50,51] Presynaptic boutons generally form along the long axis of axons and are filled with anywhere from neurotransmitter substances filled synaptic vesicles such as glutamate, acetylcholine, glycine, dopamine, serotonin, adrenalin, noradrenalin, or gamma amino butyric acid. These vesicles are often released in an activity dependent manner[49,52,53] into the small space between the pre- and post-synaptic membranes also called synaptic cleft. In the synaptic cleft, they bind and activate neurotransmitter receptors within the postsynaptic membrane.[2,23,35,54,55] In fact, the most principal function in the presynaptic buttons is the regulated release of neurotransmitter in a tightly regulated process called exocytosis which is accomplished through the fusions of synaptic vesicles with the presynaptic membrane.[23,54,56] Several lines of studies were carried out to characterize the components of the synaptic vesicle membrane and found a number of proteins, including SYP, synaptotagmin, and synaptobrevin that functions as the regulators of exocytosis.[2,52,57,58,59]

SYP is an integral membrane glycoprotein of synaptic vesicles, with a molecular weight of 38 kDa containing four membrane-spanning domains located on the cytoplasmic surface.[2,23,60,61] It is a major component of synaptic vesicles membrane and involved in the release of neurotransmitters and formation and recycling of synaptic vesicles.[2,23,60,62,63,64] SYP was one of the first proteins to be characterized in the synaptic vesicles membrane and accounts for about 7%–10% of total synaptic vesicle proteins.[23,59,63,65,66] Based on its structure, it was proposed that SYP forms a channel in the synaptic vesicle membrane and acts as the major Ca²⁺ binding protein in synaptic vesicles. Therefore, it is suggested that SYP is involved in calcium dependent synaptic vesicle exocytosis.[60,64,67,68] In the previous investigations, SYP has also been considered as a reliable marker for synaptic density and synaptogenesis.[2,23,69,70,71,72]

The expression of SYP already starts during early neurogenesis in embryonic developing brain and is greatly up-regulated during synaptogenesis.[23,73,74] There is also evidence demonstrating that upregulation of SYP expression may contribute to the mechanisms underlying learning and memory.[75,76,77] Conversely, aberrant SYP expression has been associated with several psychiatric disorders and neurodegenerative diseases, such as, Parkinson disease, and Alzheimer's disease.[2,23,78,79,80] Earlier studies on aging and neurodegenerative disorders have correlated change in SYP immunoreactivity with loss or increase in synaptic densities.[62,76,81] In a study by Thome et al., the researchers revealed that stress exposure leads to the reduction in hippocampal expression of SYP.[82]

Cerebellum

The cerebellum is one of the most studied parts of the brain located at the back of the brain, underlying the temporal and occipital lobes of the brain.[83,84,85] For a long time, the cerebellum has been considered to be a critical brain structure for the coordination and control of voluntary movements.[2,13,16,29,86] However, recent evidence indicates that the cerebellum also plays a role in cognitive, behavioral, and emotional functions.[13,29,87] Several lines of studies clearly indicated that the cerebellum may be involved in a variety of nonmotor functions, including sensory discrimination, working memory, semantic association, attention, and verbal learning and memory.[88] Anyway, arranged three-layered cerebellar cortex and well-defined afferent and efferent fiber connections make it a favorite field for research on development and fiber connections of the CNS.[83,84]

The cerebellum in rats develops over a long period, extending from the early embryonic period until the first postnatal weeks. This protracted development makes it vulnerable to a broad spectrum of developmental abnormalities.[23,83,89] The development of the cerebellum occurs in four basic steps: (1) characterization of the cerebellar territory at the boundary between midbrain and hindbrain; (2) formation of two compartments for cell proliferation including the Purkinje cells which arise from the ventricular zone of the mesencephalic alar plate and granule cell precursors which formed from the upper rhombic lip; (3) migration of the granule cells: granule precursor cells form the EGL, from which granule cells migrate inward to their definite position in the IGL, and (4) formation of cerebellar circuitry and further differentiation.[90,91,92,93,94,95]

This phenomenon in the developing rat cerebellar cortex has been studied in detail by Altman and Bayer.[90,96,97] Immediately after birth, granule cells are observed to be aligned outside of the developing cerebellum, known as EGL. At the next postnatal days, the granule cells extrude processes that move toward Purkinje cells. Finally, the cell bodies of the granule cells pass through the layer of Purkinje cells to form IGL. At the same time, Purkinje cells also develop dendrites, and begin to form synapses with parallel fibers, and form the ML. Approximately 2 weeks after birth, formation of the ML is completed, and the cerebellum shows almost the same morphology as that of the adult rat.[13,23,90,96,97]

Effects of maternal diabetes in pregnancy on synaptogenesis in developing cerebellum

The adverse impacts of diabetes during pregnancy on the developing CNS of the fetuses and newborns are already well documented.[6,7,17,98,99,100,101] However, there is limited number of studies that specially focused on the impacts of diabetes during pregnancy on synaptogenesis in the developing cerebellum of offspring. The earlier investigations demonstrated a moderate disturbance in memory and learning and complex information processing of offspring born to mothers who had diabetes during their pregnancy.[102,103] Rizzo et al., in their investigation demonstrated a significant correlation between gestational diabetes and lower IQ in children.[104]

Other studies also revealed a negative relationship between the severity of maternal hyperglycemia during and the gestation and the performance of the children on various developmental and behavioral tests.[7,19,105,106,107] A research by Ornoy et al. reported that children younger than 9 years, born to diabetic women, had a higher rate of attention deficit, lower cognitive scores, and lower gross and fine motor achievements.[108] In another study, Rizzo et al. revealed a striking relationship between second- and third-trimester glycemic control and poorer infant responses on the Brazelton Neonatal Behavioral Assessment Scale.[104] Taken together, the existing evidence not only demonstrate the teratogenic effect of maternal diabetes on the development of offspring's nervous system but also provide the perhaps earliest indicator of postnatal CNS problems reflected in intellectual, behavioral and movement anomalies exhibited by children of mothers with diabetes.[7,18,27,28,98,107] Nevertheless, no report can fully explore the molecular mechanisms of maternal diabetes-induced neurodevelopmental and neurofunctional abnormalities.[2,7,23]

In a recent study by Hami et al., the authors evaluated the effects of diabetes in pregnancy on the expression and localization of SYP during the development of cerebellum in the rat offspring. The authors found no significant alterations in the cerebellar expression/localization of SYP in neonate's born to diabetic animals, immediately after birth. Nevertheless, they reported that the expression of SYP was significantly down-regulated at 1- and 2-weeks-old of age rats. In addition, their results also demonstrated that the localization of SYP protein was strikingly reduced in all three distinct layers of cerebellar cortex of neonates born to diabetic animals, especially at postnatal day (P) 14.[23] Another recent study by Vafaei-Nezhad et al. indicated that there were no differences in the SYP expression/localization in the hippocampus of neonates born to diabetic dams at P0; the researchers also indicated that the gestational diabetes in pregnancy is associated with a significant downregulation in hippocampal expression/localization of SYP in the neonatal rats at P7, and P14.[2] Since SYP and other synaptic vesicle proteins have been implicated in the mechanisms of cellular plasticity underlying learning;[62,75,76,77,81] the early decrease in SYP expression/localization may reflect a downregulation of synaptic function and may be related to the reduction in synaptic density and might disrupt the development and function of cerebellum.[23,76,77,79,109]

Taken together, The exact mechanisms through which diabetes in pregnancy affects fetal CNS development and function are not completely understood, the conclusion from a review of numerous studies is that maternal hyperglycemia during pregnancy may be a major teratogenic factor.[7,13,31] The remarkable thing about the glucose, in contrast to insulin, is that Glucose can freely cross the placental barrier; thus, maternal hyperglycemia during gestational period causes fetal hyperglycemia.[7,23,110,111] Fetal hyperglycemia stimulates the developing,[112] resulting to in utero fetal hyperinsulinemia. On separation of the newborn from the mother, the glucose former no longer is supported by placental glucose transfer, may develop neonatal hypoglycemia.[7,15]

In addition, some researchers argue that the maternal metabolic condition, in combination with a disturbed fetal metabolism, may have teratogenic consequences and may increase the risk of fetal abnormalities in diabetic pregnancies.[24,104] Excess fetal reactive oxygen species have also been implicated in the pathogenesis of diabetes-induced congenital anomalies in CNS.[113,114,115,116]

CONCLUSION

In these studies using SYP, a marker of synaptic density and synaptic vesicle formation, it was shown that maternal hyperglycemia, in combination with neonatal hyperinsulinemia was able to decline synaptogenesis in the offspring's cerebellar cortex. This alteration may result in a delay in normal cerebellar development and function and could be a reason for the structural, behavioral, and cognitive abnormalities observed in the offspring of diabetic mothers.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.