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Correlation between local monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) densities in the adult rat brain
Neuroscience Letters 355 (2004) 105–108 www.elsevier.com/locate/neulet
Correlation between local monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) densities in the adult rat brain Martin H. Maurer, Martin Canis, Wolfgang Kuschinsky*, Roman Duelli Department of Physiology and Pathophysiology, University of Heidelberg, Im Neuenheimer Feld 326, 69120 Heidelberg, Germany Received 14 August 2003; received in revised form 22 October 2003; accepted 23 October 2003
Abstract Monocarboxylate transporters type 1 (MCT1) facilitate the transport of monocarboxylates across cell membranes of the blood – brain barrier and brain parenchymal cells. The present study had two aims: (1) to determine the local distribution of MCT1 in the brain; and (2) to compare the local densities of MCT1 with the local densities of the main nutritional transporters, glucose transporter GLUT1. Using immunoautoradiography of cryosections from rat brain, 32 brain structures were analyzed. (1) A heterogenous distribution pattern of MCT1 densities was observed throughout the brain. Compared to brain homogenate (100%), MCT1 densities ranged from 43 to 164% in the brain structures investigated. Local GLUT1 densities showed a comparable range (35– 145%). (2) A close correlation was found between local MCT1 and local GLUT1 densities. As local GLUT1 densities reflect local glucose metabolism in the brain, we conclude that local MCT1 densities are adjusted to local glucose metabolism and transport. q 2003 Elsevier Ireland Ltd. All rights reserved. Keywords: Monocarboxylate transporter MCT1; Glucose transporter GLUT1; Rat brain; Autoradiography
Under physiological conditions glucose is the major substrate for brain metabolism. The transport of glucose across the blood –brain barrier and the uptake into glial and neural cells is facilitated by integral membrane proteins of the glucose transporter family [19]. Under normal conditions, alternative substrates are of minor importance for the energy metabolism. Monocarboxylates such as beta-hydroxybutyrate, acetoacetate, lactate, and pyruvate become relevant during the neonatal period, long term fasts, or high fat diet [9,12,13], since they can substitute for glucose. Similar to the transport of glucose, the transport of monocarboxylates is facilitated by specialized transporters. Several isoforms of these monocarboxylate transporters (MCTs) are expressed in the brain [5 –7,14,17]. As the most important monocarboxylate transporter, MCT1 is mainly expressed in endothelial cells and at the tissue-cerebrospinal fluid interface including ependymocytes, glial limiting membranes, and choroid plexus epithelium [5]. Its expression is also detectable in astrocytes and neurons [5,14]. * Corresponding author. Tel.: þ49-6221-54-4033; fax: þ 49-6221-544561. E-mail address: [email protected] (W. Kuschinsky).
Glucose transporters (GLUTs) and MCTs facilitate the transport of the nutrients into the brain tissue, thus enabling a sufficient delivery of substrates for metabolism into the brain tissue. As to glucose metabolism, it is well known that glucose utilization is distributed heterogeneously in the brain [18]. Accordingly, a heterogenous distribution of glucose transporters GLUT1 and GLUT3 has been shown in the brain which corresponds to the distribution of glucose utilization [2,3]. In contrast, little is known about the local densities of MCT1 throughout the brain. Immunhistochemical staining techniques have demonstrated a widespread distribution of MCT1 in adult rat brains although the local densities of MCT1 were not determined. Therefore, the present study had the aim to investigate the local distribution of MCT1 in different brain structures. To this end, a modified autoradiographic technique was developed which allows the analysis of the local distribution of MCT1 from autoradiograms of cryosections of rat brains. For comparison, local glucose transporter GLUT1 densities were also determined in the same brain structures. Experiments were performed on eight adult Sprague– Dawley rats weighing 250– 350 g. Protocols are concordant with the policy on the use of animals, as endorsed by the National Research Council of the USA, and fulfil the
0304-3940/03/$ - see front matter q 2003 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.neulet.2003.10.056
106
M.H. Maurer et al. / Neuroscience Letters 355 (2004) 105–108
requirements of German law (approved by Regierungspra¨sidium Karlsruhe). Briefly, animals were anesthesized by isoflurane inhalation and sacrificed by decapitation. Brains were removed rapidly and shock-frozen in 2-methylbutane (Merck, Darmstadt, Germany), chilled to 2 40 8C and embedded in M-1 embedding matrix (Lipshaw, Detroit, MI, USA). Coronal cryosections of 10 mm thickness were placed on polylysine-coated slides. Air-dried sections were fixed with absolute acetone at room temperature for 2 min and transferred to 0.2% Triton X-100 (Sigma, Deisenhofen, Germany) in phosphate buffered saline (PBS). For MCT1 immunostaining, sections were incubated overnight at 4 8C in a humified incubation chamber with a chicken polyclonal antibody directed against a synthetic 13 amino acid C-terminal sequence peptide of MCT1 (Chemicon, Temecula, CA, USA) diluted 1:125 in PBS to provide epitope saturation. The next day, sections were washed 3 £ 5 min in 0.2% Tween-20 (Sigma, Deisenhofen, Germany) in PBS and incubated with a rabbit-anti-chicken secondary adapter antibody (Jackson Immuno Research Labaratories, West Grove, PA, USA) for 60 min, diluted 1:375 in PBS at saturation. Slides were washed 3 £ 5 min in 0.2% Tween-20 in PBS and incubated with a radiolabeled antibody ([35S]-anti-rabbit IgG, Amersham, Braunschweig, Germany) for 30 min, with a specific activity of 0.24 Ci/mM. Slides were rinsed 3 £ 5 min in 0.2% Tween-20 in PBS, dehydrated and exposed to Kodak MinR1 X-ray films for 3 weeks. We measured local tissue concentrations of [35S] by densitometry of the autoradiograms with an image analyzing system (MCID, Imaging Research Inc., St. Catherines, Ont., Canada). Values determined for each brain structure represent a mean of at least six measurements of optical density for that structure in three consecutive sections. Measured values were calibrated by comparison to a co-exposed [14C] standard and normalized by immunostained brain homogenate which has been made by homogenization of five rat brains. [14C] standards were used because of the long half life of [14C]. For antibody tagging [14C] tagged antibodies were not available. [35S] was therefore used since its radiation characteristics are close to those of [14C]. [14C] standards were calibrated for [35S] using brain paste loaded with [35S]. All values are means ^ SD. Omission of the primary antibody, or incubation with preimmune rabbit serum, produced faint or no staining. The results obtained for MCT1 densities were finally compared to previously reported GLUT1 densities [2 – 4]. To determine the local distribution of MCT1 in the rat brain, 32 brain structures were investigated by immunoautoradiography. Measured optical densities of MCT1 in the structures investigated ranged from 43 to 164% when compared to brain homogenate as 100% (Table 1). Lowest values were obtained in the internal capsule, followed by the two white matter structures corpus callosum and cerebral peduncle. A typical autoradiogram of the distribution of MCT1 is shown in Fig. 1A for a brain coronal section. The
Table 1 Regional distribution of monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) densities in the adult rat brain Structure
MCT1 (mean ^ SD)
GLUT1 (mean ^ SD)
Internal capsule Cerebral peduncle Corpus callosum, genu corpus callosum, forceps major Cerebellar peduncule Deep mesencephalic nucleus Entorhinal cortex Amygdala Caudate nucleus Hippocampus, CA2 Substantia nigra, compact Central grey Dentate gyrus Hippocampus, CA1 Superior colliculus, all layers Lateral geniculate body, ventral Thalamus, ventroposterior nucleus Retrosplenial cortex Thalamus, lateral posterior nucleus Substantia nigra, reticular Superior colliculus, layers 4–6 Cingulate cortex Posterior hypothalamic nucleus Medial geniculate body Auditory cortex Occipital cortex, area 18 Lateral geniculate body, dorsal Hippocampus, lateral ependyma Frontoparietal cortex Superior colliculus, layers 1–3 Mammilary body Hippocampus, medial ependyma Brain homogenate
43 ^ 5 49 ^ 5 51 ^ 6 53 ^ 6 62 ^ 5 82 ^ 10 94 ^ 7 94 ^ 3 96 ^ 5 99 ^ 5 100 ^ 9 101 ^ 9 103 ^ 9 103 ^ 12 105 ^ 10 105 ^ 4 106 ^ 4 107 ^ 7 107 ^ 4 109 ^ 11 110 ^ 11 111 ^ 6 113 ^ 8 114 ^ 9 119 ^ 6 121 ^ 9 121 ^ 8 121 ^ 11 121 ^ 5 137 ^ 14 153 ^ 20 164 ^ 15 100 ^ 3
57 ^ 7 54 ^ 9 39 ^ 2 63 ^ 11 35 ^ 5 85 ^ 12 97 ^ 12 121 ^ 9 126 ^ 10 114 ^ 13 110 ^ 14 108 ^ 25 117 ^ 13 86 ^ 15 108 ^ 23 93 ^ 12 131 ^ 16 122 ^ 21 128 ^ 8 133 ^ 20 88 ^ 13 97 ^ 11 109 ^ 13 131 ^ 19 133 ^ 17 120 ^ 20 105 ^ 7 93 ^ 13 145 ^ 20 111 ^ 16 132 ^ 19 145 ^ 11 100 ^ 7
Local densities were normalized to brain homogenate ( ¼ 100%).
corresponding autoradiogram for GLUT1 is shown in Fig. 1B. The optical density is proportional to the density of MCT1 protein. This holds for all brain structures. The heterogeneous distribution of MCT1 is evident. Highest MCT1 densities were found in the mammilary body and the hippocampus. MCT1 densities in each brain structure were compared to GLUT1 densities in the same brain structure. A close correlation was found between the densities of MCT1 and GLUT1 (R ¼ 0:80) (Fig. 2). Blood-borne monocarboxylates have to be transported through the membranes of brain endothelial and parenchymal cells in order to reach their site of utilization in the brain [15]. The monocarboxylic acid transporter MCT1 is known to be expressed in endothelial cells of the blood – brain barrier [5]. The autoradiographic method used in the present study does not allow to differentiate between different cellular locations of MCT1. Therefore only heterogeneities far beyond the cellular level can be detected. MCT1 densities have been found to be increased after diet-induced ketosis [10]. This increase in MCT1 densities
M.H. Maurer et al. / Neuroscience Letters 355 (2004) 105–108
during chronic ketosis induced by a ketogenic diet has been linked to the concept that the brain can partially replace glucose by ketone bodies under certain circumstances as major metabolic source of energy [16]. The densities of MCT1 seem to be dependent on the availability of its substrate, thus MCT1 densities should be increased when the supply of lactate is augmented. In the suckling period, nutrition contains larger amounts of lactate than glucose, and indeed, increased MCT1 densities have been found in 17-day old suckling rats, compared to adult animals [9]. In the suckling rats, MCT1 densities were increased by factor 19 in astrocytic membranes. These results support the hypothesis that lactate could also contribute to the brain cell energy metabolism under certain conditions in a complimentary way to glucose [11]. Patients with glucose transporter protein syndrome [1] benefit from a ketogenic diet [8], if this syndrome is diagnosed early in development. In this disease, blood – brain glucose transport is severely impaired because of GLUT1 mutations resulting in dysfunctional transporters resulting in mental retardation. The dietary benefit occurs
107
Fig. 2. Correlation between local densities of monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) in the adult rat brain. The correlation coefficient is R ¼ 0:80, indicating a close relationship between both parameters.
because increased plasma ketone body concentrations supplement the deficient glucose supply and provide a carbon source for precursors of essential lipid and protein biosynthesis associated with brain growth and development. Our finding that the MCT1 protein is regulated according to the local demands of the metabolism make it likely that, in addition to the increased ketone body concentration, the upregulation of MCT1 may play a role in the dietary amelioration of this syndrome. In the present study, local densities of MCT1 showed a close correlation with the densities of GLUT1 in the different brain structures investigated, indicating that the density of MCT1 is related to the density of GLUT1 and local cerebral glucose utilization [4]. Thus, brain regions with high glucose uptake and utilization are also able to transport increased amounts of monocarboxylates. In summary, the findings of the present study demonstrate a close correlation between MCT1 and GLUT1 densities in the brain of adult rats, indicating a parallel capacity of glucose and monocarboxylate transport in different brain structures.
References
Fig. 1. Typical autoradiograms of brain sections exposed to antibodies against: (A) monocarboxylate transporter 1 (MCT1); and (B) glucose transporter 1 (GLUT1). Optical densities correspond to transporter densities, a high optical density represents a high transporter density and vice versa.
[1] D.C. De Vivo, R.R. Trifiletti, R.I. Jacobson, G.M. Ronen, R.A. Behmand, S.I. Harik, Defective glucose transport across the blood– brain barrier as a cause of persistent hypoglycorrhachia, seizures, and developmental delay, N. Engl. J. Med. 325 (1991) 703–709. [2] R. Duelli, M.H. Maurer, S. Heiland, V. Elste, W. Kuschinsky, Brain water content, glucose transporter densities and glucose utilization after 3 days of water deprivation in the rat, Neurosci. Lett. 271 (1999) 13–16. [3] R. Duelli, M.H. Maurer, W. Kuschinsky, Decreased glucose transporter densities, rate constants and glucose utilization in visual structures of rat brain during chronic visual deprivation, Neurosci. Lett. 250 (1998) 49 –52. [4] R. Duelli, M.H. Maurer, R. Staudt, S. Heiland, L. Duembgen, W.
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M.H. Maurer et al. / Neuroscience Letters 355 (2004) 105–108 Kuschinsky, Increased cerebral glucose utilization and decreased glucose transporter Glut1 during chronic hyperglycemia in rat brain, Brain Res. 858 (2000) 338–347. D.Z. Gerhart, B.E. Enerson, O.Y. Zhdankina, R.L. Leino, L.R. Drewes, Expression of monocarboxylate transporter MCT1 by brain endothelium and glia in adult and suckling rats, Am. J. Physiol. 273 (1997) E207– E213. D.Z. Gerhart, B.E. Enerson, O.Y. Zhdankina, R.L. Leino, L.R. Drewes, Expression of the monocarboxylate transporter MCT2 by rat brain glia, Glia 22 (1998) 272– 281. A.P. Halestrap, N.T. Price, The proton-linked monocarboxylate transporter (MCT) family: structure, function and regulation, Biochem. J. 343 (1999) 281–299. J. Klepper, D. Wang, J. Fischbarg, J.C. Vera, I.T. Jarjour, K.R. O’Driscoll, D.C. De Vivo, Defective glucose transport across brain tissue barriers: a newly recognized neurological syndrome, Neurochem. Res. 24 (1999) 587 –594. R.L. Leino, D.Z. Gerhart, L.R. Drewes, Monocarboxylate transporter (MCT1) abundance in brains of suckling and adult rats: a quantitative electron microscopic immunogold study, Brain Res. Dev. Brain Res. 113 (1999) 47 –54. R.L. Leino, D.Z. Gerhart, R. Duelli, B.E. Enerson, L.R. Drewes, Dietinduced ketosis increases monocarboxylate transporter (MCT1) levels in rat brain, Neurochem. Int. 38 (2001) 519–527. P.J. Magistretti, L. Pellerin, D.L. Rothman, R.G. Shulman, Energy on demand, Science 283 (1999) 496 –497. G.A. Mitchell, S. Kassovska-Bratinova, Y. Boukaftane, M.F. Robert, S.P. Wang, L. Ashmarina, M. Lambert, P. Lapierre, E. Potier, Medical
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aspects of ketone body metabolism, Clin. Invest. Med. 18 (1995) 193 –216. A. Nehlig, A. Pereira de Vasconcelos, Glucose and ketone body utilization by the brain of neonatal rats, Prog. Neurobiol. 40 (1993) 163 –221. L. Pellerin, G. Pellegri, P.G. Bittar, Y. Charnay, C. Bouras, J.L. Martin, N. Stella, P.J. Magistretti, Evidence supporting the existence of an activity-dependent astrocyte-neuron lactate shuttle, Dev. Neurosci. 20 (1998) 291 –299. L. Pellerin, G. Pellegri, J.L. Martin, P.J. Magistretti, Expression of monocarboxylate transporter mRNAs in mouse brain: support for a distinct role of lactate as an energy substrate for the neonatal vs. adult brain, Proc. Natl. Acad. Sci. USA 95 (1998) 3990–3995. K. Pierre, L. Pellerin, R. Debernardi, B.M. Riederer, P.J. Magistretti, Cell-specific localization of monocarboxylate transporters, MCT1 and MCT2, in the adult mouse brain revealed by double immunohistochemical labeling and confocal microscopy, Neuroscience 100 (2000) 617 –627. N.T. Price, V.N. Jackson, A.P. Halestrap, Cloning and sequencing of four new mammalian monocarboxylate transporter (MCT) homologues confirms the existence of a transporter family with an ancient past, Biochem. J. 329 (1998) 321–328. L. Sokoloff, M. Reivich, C. Kennedy, M.H. Des-Rosiers, C.S. Patlak, K.D. Pettigrew, O. Sakurada, M. Shinohara, The [14C]deoxyglucose method for the measurement of local cerebral glucose utilization: theory, procedure, and normal values in the conscious and anesthetized albino rat, J. Neurochem. 28 (1977) 897–916. S.J. Vannucci, F. Maher, I.A. Simpson, Glucose transporter proteins in brain: delivery of glucose to neurons and glia, Glia 21 (1997) 2–21.
Correlation between local monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) densities in the adult rat brain Martin H. Maurer, Martin Canis, Wolfgang Kuschinsky*, Roman Duelli Department of Physiology and Pathophysiology, University of Heidelberg, Im Neuenheimer Feld 326, 69120 Heidelberg, Germany Received 14 August 2003; received in revised form 22 October 2003; accepted 23 October 2003
Abstract Monocarboxylate transporters type 1 (MCT1) facilitate the transport of monocarboxylates across cell membranes of the blood – brain barrier and brain parenchymal cells. The present study had two aims: (1) to determine the local distribution of MCT1 in the brain; and (2) to compare the local densities of MCT1 with the local densities of the main nutritional transporters, glucose transporter GLUT1. Using immunoautoradiography of cryosections from rat brain, 32 brain structures were analyzed. (1) A heterogenous distribution pattern of MCT1 densities was observed throughout the brain. Compared to brain homogenate (100%), MCT1 densities ranged from 43 to 164% in the brain structures investigated. Local GLUT1 densities showed a comparable range (35– 145%). (2) A close correlation was found between local MCT1 and local GLUT1 densities. As local GLUT1 densities reflect local glucose metabolism in the brain, we conclude that local MCT1 densities are adjusted to local glucose metabolism and transport. q 2003 Elsevier Ireland Ltd. All rights reserved. Keywords: Monocarboxylate transporter MCT1; Glucose transporter GLUT1; Rat brain; Autoradiography
Under physiological conditions glucose is the major substrate for brain metabolism. The transport of glucose across the blood –brain barrier and the uptake into glial and neural cells is facilitated by integral membrane proteins of the glucose transporter family [19]. Under normal conditions, alternative substrates are of minor importance for the energy metabolism. Monocarboxylates such as beta-hydroxybutyrate, acetoacetate, lactate, and pyruvate become relevant during the neonatal period, long term fasts, or high fat diet [9,12,13], since they can substitute for glucose. Similar to the transport of glucose, the transport of monocarboxylates is facilitated by specialized transporters. Several isoforms of these monocarboxylate transporters (MCTs) are expressed in the brain [5 –7,14,17]. As the most important monocarboxylate transporter, MCT1 is mainly expressed in endothelial cells and at the tissue-cerebrospinal fluid interface including ependymocytes, glial limiting membranes, and choroid plexus epithelium [5]. Its expression is also detectable in astrocytes and neurons [5,14]. * Corresponding author. Tel.: þ49-6221-54-4033; fax: þ 49-6221-544561. E-mail address: [email protected] (W. Kuschinsky).
Glucose transporters (GLUTs) and MCTs facilitate the transport of the nutrients into the brain tissue, thus enabling a sufficient delivery of substrates for metabolism into the brain tissue. As to glucose metabolism, it is well known that glucose utilization is distributed heterogeneously in the brain [18]. Accordingly, a heterogenous distribution of glucose transporters GLUT1 and GLUT3 has been shown in the brain which corresponds to the distribution of glucose utilization [2,3]. In contrast, little is known about the local densities of MCT1 throughout the brain. Immunhistochemical staining techniques have demonstrated a widespread distribution of MCT1 in adult rat brains although the local densities of MCT1 were not determined. Therefore, the present study had the aim to investigate the local distribution of MCT1 in different brain structures. To this end, a modified autoradiographic technique was developed which allows the analysis of the local distribution of MCT1 from autoradiograms of cryosections of rat brains. For comparison, local glucose transporter GLUT1 densities were also determined in the same brain structures. Experiments were performed on eight adult Sprague– Dawley rats weighing 250– 350 g. Protocols are concordant with the policy on the use of animals, as endorsed by the National Research Council of the USA, and fulfil the
0304-3940/03/$ - see front matter q 2003 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.neulet.2003.10.056
106
M.H. Maurer et al. / Neuroscience Letters 355 (2004) 105–108
requirements of German law (approved by Regierungspra¨sidium Karlsruhe). Briefly, animals were anesthesized by isoflurane inhalation and sacrificed by decapitation. Brains were removed rapidly and shock-frozen in 2-methylbutane (Merck, Darmstadt, Germany), chilled to 2 40 8C and embedded in M-1 embedding matrix (Lipshaw, Detroit, MI, USA). Coronal cryosections of 10 mm thickness were placed on polylysine-coated slides. Air-dried sections were fixed with absolute acetone at room temperature for 2 min and transferred to 0.2% Triton X-100 (Sigma, Deisenhofen, Germany) in phosphate buffered saline (PBS). For MCT1 immunostaining, sections were incubated overnight at 4 8C in a humified incubation chamber with a chicken polyclonal antibody directed against a synthetic 13 amino acid C-terminal sequence peptide of MCT1 (Chemicon, Temecula, CA, USA) diluted 1:125 in PBS to provide epitope saturation. The next day, sections were washed 3 £ 5 min in 0.2% Tween-20 (Sigma, Deisenhofen, Germany) in PBS and incubated with a rabbit-anti-chicken secondary adapter antibody (Jackson Immuno Research Labaratories, West Grove, PA, USA) for 60 min, diluted 1:375 in PBS at saturation. Slides were washed 3 £ 5 min in 0.2% Tween-20 in PBS and incubated with a radiolabeled antibody ([35S]-anti-rabbit IgG, Amersham, Braunschweig, Germany) for 30 min, with a specific activity of 0.24 Ci/mM. Slides were rinsed 3 £ 5 min in 0.2% Tween-20 in PBS, dehydrated and exposed to Kodak MinR1 X-ray films for 3 weeks. We measured local tissue concentrations of [35S] by densitometry of the autoradiograms with an image analyzing system (MCID, Imaging Research Inc., St. Catherines, Ont., Canada). Values determined for each brain structure represent a mean of at least six measurements of optical density for that structure in three consecutive sections. Measured values were calibrated by comparison to a co-exposed [14C] standard and normalized by immunostained brain homogenate which has been made by homogenization of five rat brains. [14C] standards were used because of the long half life of [14C]. For antibody tagging [14C] tagged antibodies were not available. [35S] was therefore used since its radiation characteristics are close to those of [14C]. [14C] standards were calibrated for [35S] using brain paste loaded with [35S]. All values are means ^ SD. Omission of the primary antibody, or incubation with preimmune rabbit serum, produced faint or no staining. The results obtained for MCT1 densities were finally compared to previously reported GLUT1 densities [2 – 4]. To determine the local distribution of MCT1 in the rat brain, 32 brain structures were investigated by immunoautoradiography. Measured optical densities of MCT1 in the structures investigated ranged from 43 to 164% when compared to brain homogenate as 100% (Table 1). Lowest values were obtained in the internal capsule, followed by the two white matter structures corpus callosum and cerebral peduncle. A typical autoradiogram of the distribution of MCT1 is shown in Fig. 1A for a brain coronal section. The
Table 1 Regional distribution of monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) densities in the adult rat brain Structure
MCT1 (mean ^ SD)
GLUT1 (mean ^ SD)
Internal capsule Cerebral peduncle Corpus callosum, genu corpus callosum, forceps major Cerebellar peduncule Deep mesencephalic nucleus Entorhinal cortex Amygdala Caudate nucleus Hippocampus, CA2 Substantia nigra, compact Central grey Dentate gyrus Hippocampus, CA1 Superior colliculus, all layers Lateral geniculate body, ventral Thalamus, ventroposterior nucleus Retrosplenial cortex Thalamus, lateral posterior nucleus Substantia nigra, reticular Superior colliculus, layers 4–6 Cingulate cortex Posterior hypothalamic nucleus Medial geniculate body Auditory cortex Occipital cortex, area 18 Lateral geniculate body, dorsal Hippocampus, lateral ependyma Frontoparietal cortex Superior colliculus, layers 1–3 Mammilary body Hippocampus, medial ependyma Brain homogenate
43 ^ 5 49 ^ 5 51 ^ 6 53 ^ 6 62 ^ 5 82 ^ 10 94 ^ 7 94 ^ 3 96 ^ 5 99 ^ 5 100 ^ 9 101 ^ 9 103 ^ 9 103 ^ 12 105 ^ 10 105 ^ 4 106 ^ 4 107 ^ 7 107 ^ 4 109 ^ 11 110 ^ 11 111 ^ 6 113 ^ 8 114 ^ 9 119 ^ 6 121 ^ 9 121 ^ 8 121 ^ 11 121 ^ 5 137 ^ 14 153 ^ 20 164 ^ 15 100 ^ 3
57 ^ 7 54 ^ 9 39 ^ 2 63 ^ 11 35 ^ 5 85 ^ 12 97 ^ 12 121 ^ 9 126 ^ 10 114 ^ 13 110 ^ 14 108 ^ 25 117 ^ 13 86 ^ 15 108 ^ 23 93 ^ 12 131 ^ 16 122 ^ 21 128 ^ 8 133 ^ 20 88 ^ 13 97 ^ 11 109 ^ 13 131 ^ 19 133 ^ 17 120 ^ 20 105 ^ 7 93 ^ 13 145 ^ 20 111 ^ 16 132 ^ 19 145 ^ 11 100 ^ 7
Local densities were normalized to brain homogenate ( ¼ 100%).
corresponding autoradiogram for GLUT1 is shown in Fig. 1B. The optical density is proportional to the density of MCT1 protein. This holds for all brain structures. The heterogeneous distribution of MCT1 is evident. Highest MCT1 densities were found in the mammilary body and the hippocampus. MCT1 densities in each brain structure were compared to GLUT1 densities in the same brain structure. A close correlation was found between the densities of MCT1 and GLUT1 (R ¼ 0:80) (Fig. 2). Blood-borne monocarboxylates have to be transported through the membranes of brain endothelial and parenchymal cells in order to reach their site of utilization in the brain [15]. The monocarboxylic acid transporter MCT1 is known to be expressed in endothelial cells of the blood – brain barrier [5]. The autoradiographic method used in the present study does not allow to differentiate between different cellular locations of MCT1. Therefore only heterogeneities far beyond the cellular level can be detected. MCT1 densities have been found to be increased after diet-induced ketosis [10]. This increase in MCT1 densities
M.H. Maurer et al. / Neuroscience Letters 355 (2004) 105–108
during chronic ketosis induced by a ketogenic diet has been linked to the concept that the brain can partially replace glucose by ketone bodies under certain circumstances as major metabolic source of energy [16]. The densities of MCT1 seem to be dependent on the availability of its substrate, thus MCT1 densities should be increased when the supply of lactate is augmented. In the suckling period, nutrition contains larger amounts of lactate than glucose, and indeed, increased MCT1 densities have been found in 17-day old suckling rats, compared to adult animals [9]. In the suckling rats, MCT1 densities were increased by factor 19 in astrocytic membranes. These results support the hypothesis that lactate could also contribute to the brain cell energy metabolism under certain conditions in a complimentary way to glucose [11]. Patients with glucose transporter protein syndrome [1] benefit from a ketogenic diet [8], if this syndrome is diagnosed early in development. In this disease, blood – brain glucose transport is severely impaired because of GLUT1 mutations resulting in dysfunctional transporters resulting in mental retardation. The dietary benefit occurs
107
Fig. 2. Correlation between local densities of monocarboxylate transporter 1 (MCT1) and glucose transporter 1 (GLUT1) in the adult rat brain. The correlation coefficient is R ¼ 0:80, indicating a close relationship between both parameters.
because increased plasma ketone body concentrations supplement the deficient glucose supply and provide a carbon source for precursors of essential lipid and protein biosynthesis associated with brain growth and development. Our finding that the MCT1 protein is regulated according to the local demands of the metabolism make it likely that, in addition to the increased ketone body concentration, the upregulation of MCT1 may play a role in the dietary amelioration of this syndrome. In the present study, local densities of MCT1 showed a close correlation with the densities of GLUT1 in the different brain structures investigated, indicating that the density of MCT1 is related to the density of GLUT1 and local cerebral glucose utilization [4]. Thus, brain regions with high glucose uptake and utilization are also able to transport increased amounts of monocarboxylates. In summary, the findings of the present study demonstrate a close correlation between MCT1 and GLUT1 densities in the brain of adult rats, indicating a parallel capacity of glucose and monocarboxylate transport in different brain structures.
References
Fig. 1. Typical autoradiograms of brain sections exposed to antibodies against: (A) monocarboxylate transporter 1 (MCT1); and (B) glucose transporter 1 (GLUT1). Optical densities correspond to transporter densities, a high optical density represents a high transporter density and vice versa.
[1] D.C. De Vivo, R.R. Trifiletti, R.I. Jacobson, G.M. Ronen, R.A. Behmand, S.I. Harik, Defective glucose transport across the blood– brain barrier as a cause of persistent hypoglycorrhachia, seizures, and developmental delay, N. Engl. J. Med. 325 (1991) 703–709. [2] R. Duelli, M.H. Maurer, S. Heiland, V. Elste, W. Kuschinsky, Brain water content, glucose transporter densities and glucose utilization after 3 days of water deprivation in the rat, Neurosci. Lett. 271 (1999) 13–16. [3] R. Duelli, M.H. Maurer, W. Kuschinsky, Decreased glucose transporter densities, rate constants and glucose utilization in visual structures of rat brain during chronic visual deprivation, Neurosci. Lett. 250 (1998) 49 –52. [4] R. Duelli, M.H. Maurer, R. Staudt, S. Heiland, L. Duembgen, W.
108
[5]
[6]
[7]
[8]
[9]
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