Taurine and Brain Development: Trophic or Cytoprotective Actions?
Herminia Pasantes-Morales • Reyna Herna´ndez-Benı´tez
Accepted: 1 September 2010 / Published online: 15 September 2010 ti Springer Science+Business Media, LLC 2010
Abstract The decline of taurine content during brain maturation as well as the consequences of taurine defi- ciency disturbing brain development, suggest its involve- ment in basic processes of developing brain cells. If taurine participates in cell protection, differentiation or prolifera- tion in the developing brain is as yet unclear. Extensive and solid evidence supports taurine cytoprotective actions, directly or indirectly related to an antioxidant effect. Since redox status and oxidative stress are now implicated in signalling processes regulating cell differentiation and proliferation, the question is raised of whether the taurine antioxidant activity is on the basis of its requirement during brain development.
Keywords Neural precursor cells ti Oxidative stress Antioxidants ti Neurospheres ti
Taurine in Cell Physiology
Taurine is a sulphur amino acid, present in mM concen- trations in most animal cell types. Some tissues such as skeletal muscle, heart, brain and retina contain remarkably high taurine levels, often up to 40 mM. The cell taurine pool results from the concerted operation of synthesis, influx and efflux. The contribution of these processes varies
among cell types and species. In general taurine has a small turnover, particularly in those tissues with the highest taurine levels. Taurine is mostly found soluble in the cytosol. It is not a protein constituent and participates in very few cell metabolic reactions. Taurocholic acid for- mation is one of the few cell reactions in which taurine is implicated . Taurine is formed from cysteine through the sequential action of the cysteine dioxygenase and the cysteine sulfinate decarboxylase. The decarboxylase is the rate-limiting of the pathway, and its functional expression is considered as indicative of the importance of taurine synthesis in cells or tissues. In accordance with the key role of taurine in liver function via taurocholic acid formation, in most species liver is the main site of taurine synthesis. Cysteine sulfinate decarboxylase is expressed also in the mammary gland, in adipose tissue, and in low amounts in kidney, brain, lung and placenta . There is not a meta- bolic machinery for taurine degradation and the amino acid is excreted as such in urine.
Taurine formed in liver as well that coming from dietary sources is exported to other organs with lower capacities for taurine formation. This transfer occurs via an energy- Na?/Cl– dependent taurine transporter (TauT), specific for b-amino acids, with low affinity and high capacity. TauT cDNA encodes for a protein of about 70 kDa, with high homology between tissues and species. TauT is expressed in a large variety of cells and tissues, in accor- dance with the ubiquitous presence of taurine in animal cells, and shows remarkable similarities in most cells of
Special issue article in honor of Dr. Abel Lajtha. H. Pasantes-Morales (&) ti R. Herna´ndez-Benı´tez
Divisio´n de Neurociencias, Instituto de Fisiologı´a Celular, Universidad Nacional Auto´noma de Me´xico (UNAM), Ciudad Universitaria, Circuito Exterior, 04510 Mexico DF, Mexico
e-mail: [email protected]
essentially all organs. The kinetic constant of TauT in brain range for Vmax 0.39–0.67 nmoles/mg protein and for Km between 10 and 40 lM. TauT has been cloned from a variety tissues including the rat and mouse brain [3, 4].
The high taurine concentration in some tissues and its presence in most cell types suggest its involvement in a
general, basic function of the cell. Cell volume regulation seems to be this function. Maintenance of a constant vol- ume is fundamental homeostatic requirement for animal cells. Also, the volume of intracellular compartments should be maintained within strict limits to allow the proper interaction of elements of signalling chains which establish the intracellular and intercellular communication. Mechanisms devised to keep cell volume constant are present in essentially all animal cells, and have been pre- served along the evolution in most species, regardless of the constancy of their extracellular environment. Volume regulation occurs via mobilization of osmotically active solutes, osmolytes, which are present in the intracellular and extracellular compartments in amounts sufficiently large to drive water movements of the magnitude necessary to correct cell volume changes . Taurine is one of these osmolytes, and due to its special features, it is considered as an ideal osmolyte. The high taurine cell concentration together with its low or null involvement in cell metabo- lism allows taurine movements across the cell compart- ments without modifying significantly the cellular homeostasis. In this sense, taurine is perhaps the most representative of the molecules known as compatible osmolytes. Taurine mobilization outside the cell in response to an increase in cell volume occurs via a leak pathway, with high sensitivity to swelling. Once volume has been corrected, the energy-dependent carrier leads to the taurine intracellular pool replenishment. Taurine has also a crucial role in restoring cell volume after shrinkage. In this case, the adaptive mechanism is an up-regulation of the taurine transporter, increasing taurine cell content together with osmotically driven water [6, 7]. Other organic osmolytes closely involved in this process are the polialcohols myo-inositol and sorbitol.
Taurine in Brain Development
A role for taurine in brain development has been suggested since the early observations about the differences in taurine levels between the developing and the mature brain . Taurine is the free amino acid present in the highest amount in the developing, and neonatal brain, being 3–4- times higher than in the adult. This decline is a consistent feature observed among species, regardless of their dif- ferences in taurine concentration [9, 10]. The maturation related reduction in taurine content is observed in most brain regions with the exception of the olfactory bulb in which high taurine levels persists in the adult . Inter- estingly, the olfactory bulb is the region in which neuro- genesis is preserved in the adult brain. Taurine is the free amino acid present in the highest concentrations in milk, including human milk . These results suggest a
requirement for taurine when brain cells are in the process of maturation. This was supported by studies showing the consequences for brain development in taurine deficient cats . Taurine deficiency is difficult to achieve, since facing a dietary taurine restriction, animals respond by suppressing taurine excretion in urine and faeces and most important, switching to glycocholic acid while taurocholic acid synthesis is suspended. It is likely also that other amino acids replace taurine as osmolyte. All these com- pensatory responses of the organism markedly decrease taurine turnover and maintain taurine contents during long time . Cats are an exception due to their inability to form glycocholic acid and thus taurine is mobilized to liver, decreasing the taurine pool in other tissues including brain. Taurine deficiency in cats result in blindness by destruction of retinal photoreceptors and also in cardiomyopathies caused by heart taurine decrease .
In the developing brain, taurine deficiency greatly affects cell migration and organization in two brain regions containing high taurine concentrations, i.e. cerebellum and brain cortex. In cerebellum of taurine-deprived cats the most conspicuous feature is the persistence of cerebellar granule neurons in the external cell layer and the occur- rence of numerous cells in mitotic phase, whereas cell division and migration has been completed at the same age in the brain of controls or taurine-supplemented animals. Delayed migration of neuroblasts and glioblasts is also observed in the visual cortex, resulting in a disturbed pat- tern of neuronal and astrocyte distribution among the cor- tical layers. Also noteworthy is the low number of pyramidal neurons, and the poor arborisation of those present in the cortex. Disturbed morphology of the spinal cord with abnormal alignment of the dorsal root nerves is found in taurine-deficient newborn cats . Studies in monkeys fed formulas without taurine showed prominently a defective organization of cortical layers in the visual cortex . Despite the fact that these effects of taurine deficiency have been published more than three decades ago, the mechanisms of the taurine requirement for optimal nervous system maturation are as yet unknown. Two main avenues can be considered, one is that taurine is required to maintain the stability and organization of cellular and subcellular membranes, by preventing oxidative deleteri- ous reactions or directly via an effect on proliferation and maturation of developing brain cells.
Protective Actions of Taurine
The protective effects of taurine have been observed in a large number of experimental models, in vivo and in vitro, in different cells and tissues and facing a variety of injuring stimuli. Taurine protects cells from damage induced by toxic
agents such as arsenite, ozone, CCl4, manganese, Cd, acet- aminophen, methiocarb, NaF, and ammonia. Taurine has also protective actions from injury caused by peroxynitrite, ethanol, bleomycin, streptozotocin. Taurine protection has been found also in cells damaged by b-amyloid and biliru- bin. Some taurine cell protection is also reported during ischemia–reperfusion , tissue crushing , fibrosis 
and exposure to light . Cytoprotective actions of taurine have been examined in different organs, particularly lung, liver, heart, brain and kidney, but some studies also report taurine protective actions in spinal cord, urinary bladder, retinal pigment epithelium, erythrocytes, and stomach.
Oxidative stress seems to be a common condition in most models in which taurine exhibits a protective action . The detrimental effects of oxidative stress are well known. One of them is the membrane lipid peroxidation, which has profound disturbing effects on the cell function. Ion overload, cell swelling, and excessive Ca2? influx, make an autopropagated chain of deleterious effects, often with fatal consequences for the cell. Possible mechanisms for taurine protective actions include a direct effect as a free radical scavenger, or indirectly, preventing generation of reactive oxygen species and/or protecting cell mem- branes from ROS damage . Few studies support the direct action of taurine as a free radical scavenger, sug- gesting rather indirect mechanisms of protection. The old concept of taurine as ‘‘membrane stabilizer’’  is still contemporary, and may provide a common factor to the apparently dissimilar effects of taurine. It is noteworthy that taurine protective effects occur only at very high taurine levels, in the mM range, often as high as 20 mM. This requirement of high concentrations for the protective effects of taurine do not favour actions such as modulating the activity of enzymes, or modifying signalling pathways. A low-affinity interaction of taurine with phospholipids in the membrane, particularly with the charged head groups of the neutral phospholipids, may contribute to protect cell membranes . The high concentrations of taurine required for its protective actions are not present in physiological conditions at the extracellular space. However, in most pathologies or experimental conditions in which taurine shows a protective action, swelling is concurrent factor. Under these conditions, taurine is released from the usually large intracellular pool, and may reach very high concen- trations at the extracellular space, as high as required for its protective actions.
Taurine and Cell Proliferation
Studies about taurine and cell proliferation are not numerous and contain conflicting results, showing effects of taurine either increasing or decreasing cell proliferation. Taurine
increases cell proliferation in human fetal neurons , in rabbit and human retinal pigment epithelium  and in human osteoblasts . Taurine reverses the antiprolifera- tive effect of high glucose and glycosylation end products in the glomerular mesangial and in renal tubule epithelial cells, two cell types which play a central role in the pathogenesis of diabetic nephropathy [20, 21]. Taurine effects decreasing cell proliferation have been found in hepatic stellate cells , aortic smooth muscle cells  and mice osteoclasts . Some of the proliferative effects of taurine may result from indirect actions as antioxidant as suggested for its effects in mesangial and renal tubule cells [20, 21]. In these models, taurine and other antioxidants restore the antipro- liferative action of advanced glycation-end products. Tau- rine effects on cell proliferation appear mediated by increasing the phosphorylation of the MAPKs ERK1/2 .
Neural Progenitor Cells
In a recent study we found a proliferative effect of taurine in cultured embryonic progenitor cells (NPC) obtained from the mouse mesencephalon (13.5-embryonic) . NPC isolated from embryonic brain regions proliferate in vitro as spherical free-floating aggregates termed neuro- spheres . NPC in neurospheres are able to proliferate, self-renew, and further differentiate into astroglia, oligo- dendroglia and neurons. Our study showed a consistent effect of taurine increasing neurosphere size and NPC number (Fig. 1). To obtain cultures highly enriched in NPC the excised, dispersed tissue is maintained in a serum-free medium, in the presence of EGF and bFGF, which are the trophic factors sustaining growth and proliferation of NPC . Under these conditions, other cells from the dispersed tissue are not responsive to these factors and die. Conse- quently, neurosphere-forming cells are exposed to noxious agents derived from the dying cells, and presumably require protective mechanisms for survival and prolifera- tion, which may be provided in part by taurine. Results of our study on the effect of taurine in NPC did not support a cytoprotective action of taurine on NPC, but rather showed an effect increasing cell proliferation. Taurine consistently increased the number of BrdU-positive cells, in all condi- tions, but did not induce NPC proliferation nor neuro- spheres generation in the absence of the growth factors . It seems, therefore, that the effect of taurine increasing cell proliferation occurs only when the prolif- erative process has been initiated, and in this respect, it cannot be considered a proliferative agent per se, but only as a modulator of proliferation. This effect of taurine is as yet unknown, but may be related to taurine influence on antioxidant generation or directly by its antioxidant properties.
Fig. 1 Effects of taurine on neural precursor cells (NPC). Cells were obtained from the mesencephalon of 13.5 days mice. Tissue was disaggregated and cultured in a serum-free media containing epider- mal growth factor (EGF) and fibroblast growth factor (bFGF), 20 ng/mL each. This condition sustains the NPC growth while all other cells die. Within 5 days in culture, NPC form floating spheres (neurospheres). After 5 days in culture, neurospheres were disaggre- gated and their size and number of neurosphere-forming cells were measured. a Neurospheres formed in control condition (EGF ? bFGF) or b plus taurine (10 mM); photographs were visualized at
49 magnification, scale bar = 100 lm. c Number of neurosphere- forming cells grown in conditions described in a (white bar) and b (dark bar). d Proliferation of NPC in neurospheres formed over day 4 and 5 in control cultures (white bars) or plus taurine (dark bars). Proliferation in cultures was determined by BrdU incorporation assay, identifying BrdU? cells by immunocytochemistry (Details in 25). Results are expressed as percentage of BrdU? cells present in a pool of 200 cells by each condition. Bars are means ± SEM from 6 experiments, with a significant effect at P \ 0.05 (*)
There is growing evidence about a role for reactive oxygen and nitrogen species regulating cell proliferation [27–29]. Oxygen reactive species such as H2O2 appear to serve as co-stimulatory signals for cell proliferation in various cell types, including neural stem cells . The signaling chain which involves reactive oxygen species implicates ERK1/2, as well as the expression of cell cycle associated proteins such as cyclins and p21 (Cip1) [30–33]. Taurine may then have an influence on cell proliferation through antioxidant effects or an indirect effect cell via the synthesis of taurine chloramine .
All these considerations open the intriguing possibility of a convergent point for the cytoprotective and proliferative
actions of taurine. Providing evidence for this notion is a most interesting future research avenue.
Acknowledgments Work from our laboratory was supported by grants from UNAM IMPULSA-03, DGAPA IN203410 and from CONACYT 98952. This study is part of the requirements for the PhD degree in Biomedical Sciences of Reyna Herna´ndez-Benı´tez at the Universidad Nacional Auto´noma de Me´xico, with a CONACYT, Mexico fellowship.
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