Research report
Acidic amino acid accumulation by rat choroid plexus during development

https://doi.org/10.1016/S0165-3806(97)00075-8Get rights and content

Abstract

Acidic amino acid accumulation by the choroid plexuses of the lateral ventricles was investigated using 1, 2, 3 week and adult (7–10 weeks old) rats. The accumulation from both blood and CSF sides of the choroid plexuses were investigated. The uptake from blood side was studied using the bilateral in situ brain perfusion, and time-dependent uptake profiles (2, 10, 20, and 30 min) of 14C-labelled aspartate, glutamate, and NMDA were measured. [3H]Mannitol was also included in perfusion fluid as a baseline for [14C]amino acid uptake into choroidal tissue. Uptake of [14C]aspartate and [14C]glutamate declined with age, while [14C]NMDA showed no significant uptake at any age. Twenty min [3H]mannitol uptake in the 1-week-old rat was significantly greater than the adult (P<0.05). The Kin for [14C]aspartate and [14C]glutamate obtained from multiple time uptake profiles also showed reduction with development but it was greater than that for mannitol. [14C]Aspartate declined from 69.8±21.1 μl.min−1.g−1 in the neonate to 40.6±4.0 μl.min−1.g−1 in the adult (P<0.05), while glutamate showed a sharper decline from 78.9±24.2 μl.min−1.g−1 to 17.7±5.4 μl.min−1.g−1 (P<0.01). Accumulation of 14C-labelled aspartate and glutamate by the choroid plexus from CSF side was also measured using ventriculo-cisternal perfusion. The accumulation in the adult was found to be 2–3 times greater than that in the neonatal rat (P<0.05) for both amino acids. The uptake from either side was found to be saturable, stereospecific, not inhibited by neutral amino acid analogues, and shared by both aspartate and glutamate.

Introduction

In addition to acting as protein building blocks, the acidic amino acids, aspartate and glutamate, are major excitatory neurotransmitters in the central nervous system (CNS) 11, 31. Glutamate also plays an important role in the removal of ammonia and regulation of cerebral osmotic and anionic balance [32]. However, high levels of acidic amino acids in brain interstitial fluid (ISF) can cause neuronal damage 21, 30, and brain ISF levels of this group of amino acids are strictly controlled by glial cells through removal by high affinity uptake 14, 19. Furthermore, the blood–brain and blood–cerebrospinal fluid (CSF) barriers show a lower blood to tissue permeability for acidic amino acids compared to the neutral and basic amino acids 4, 5, 29, 37. The blood–brain barrier (BBB) is mainly involved in the efflux of the acidic amino acids out of the brain into blood 10, 22, 32. However, in our recent study, it was observed that the blood–CSF barrier also plays an important part in the exchange of the acidic amino acids between blood and brain, particularly during development [4]. The choroid plexuses which are a major site for the blood–CSF barrier undergo dramatic changes during postnatal development, which are both qualitative and quantitative 16, 17, 34, 35. Recently we have shown that there is greater acidic amino acid uptake into the CSF compared to the brain in neonatal rats, while in the adult uptake into the brain was more dominant [4]. The ISF surrounding the neurones and glia is in free exchange with the CSF, since the ependymal lining of the ventricles and subarachnoid space is permeable to all small molecules [9]. Therefore the level of amino acids in the CSF may have influence on levels in the brain ISF, and hence neuronal function. In this investigation the accumulation of acidic amino acids by the choroid plexus during development has been studied using two techniques: the recently developed bilateral in situ brain perfusion technique [28]for blood-to-choroid plexus transport, and the ventriculo-cisternal perfusion technique [7]for CSF to choroid plexus transport.

Section snippets

Animals and anaesthesia

Adult Wistar rats of either sex, 7–10 weeks old, were obtained from Bantin and Kingman (UK). Pregnant rats arrived 1 week before delivering and pups then used at 1, 2, and 3 weeks of age. Rats were anaesthetised i.p. with fentanyl/fluanisone (0.3 ml/kg) and midazolam hydrochloride (2 mg/kg) and heparinised (100 000 U/kg, i.p.) in accordance with Animals (Scientific Procedures) Act 1986, UK.

Perfusion mediums

For in situ brain perfusion, a protein containing Ringer plasma substitute was used. This contained (in

Time-dependent uptake

The multiple time uptake profiles of 14C-labelled aspartate, glutamate, and NMDA into choroid plexuses of 1-, 2-, 3-week-old and adult rats are shown in Fig. 1.

[3H]Mannitol, the shaded area in each graph, was used as a vascular and extracellular space marker for the choroid plexus and measured simultaneously with the amino acids. Due to rapid accumulation, the amino acid Kin values were calculated using the line of best fit over the first 20 min of perfusion. For all the ages [14C]aspartate and

Discussion

These studies have shown variations with development in the uptake of acidic amino acids from both blood and CSF sides of the lateral ventricle choroid plexuses in the rat. [3H]Mannitol uptake from the blood side was also measured to monitor changes in the extracellular and vascular space of the choroid plexus during development. This extracellular space showed a reduction with age, in magnitude similar to the developmental changes in entry of [3H]mannitol into CSF from blood 4, 28. The values

References (37)

  • J.E Preston et al.

    The steady-state amino acid fluxes across the perfused choroid plexus of the sheep

    Brain Res.

    (1990)
  • J.E Preston et al.

    The uptake of anionic and cationic amino acids by the isolated perfused sheep choroid plexus

    Brain Res.

    (1992)
  • J.E Preston et al.

    Permeability of the developing blood–brain barrier to [14C]mannitol using the rat in situ brain perfusion technique

    Dev. Brain Res.

    (1995)
  • K Seta et al.

    Cerebral amino acid uptake in vivo in newborn mice

    Brain Res.

    (1972)
  • Q.R Smith et al.

    Kinetic analysis of [36Cl]-, [22Na]- and [3H]mannitol uptake into the in vivo choroid plexus-cerebrospinal fluid brain system: ontogeny of the blood–brain and blood–CSF barriers

    Dev. Brain Res.

    (1982)
  • D.L Yudilevich et al.

    Facilitated transport of amino acids through the blood–brain barrier of the dog studied in a single capillary circulation

    Brain Res.

    (1972)
  • W.F Agnew et al.

    Protein synthesis and transport by the rat choroid plexus and ependyma

    Cell Tissue Res.

    (1980)
  • G Baños et al.

    The influx of amino acids into the brain of the rat in vivo: the essential compared with some non-essential amino acids

    Proc. Roy. Soc. Lond. B

    (1973)
  • Cited by (20)

    • Transporters of the blood-brain and blood-CSF interfaces in development and in the adult

      2013, Molecular Aspects of Medicine
      Citation Excerpt :

      It was assumed in these studies that entry was via the blood–brain barrier interface and account was not taken of possible entry via the choroid plexuses. In addition, it was not clear from these reports if the CSF and choroid plexuses had been removed prior to analysis of brain samples; any choroid plexus tissue or CSF included in the brain samples would have led to an overestimate of the contribution of blood–brain barrier transport of the amino acids into the brain, because at least some amino acids accumulate in the choroid plexuses (al-Sarraf et al., 1997a) in addition to entering the CSF directly. Separate assessment of cerebrospinal fluid in the developing brain, which would mainly reflect entry across the choroid plexuses, has been examined by Segal and colleagues (al-Sarraf et al., 1997b).

    • Homeostasis of glutamate in brain fluids: An accelerated brain-to-blood efflux of excess glutamate is produced by blood glutamate scavenging and offers protection from neuropathologies

      2009, Neuroscience
      Citation Excerpt :

      Glu transport at the blood brain barrier (BBB) has been studied by both in vitro cell uptake assays and in vivo perfusion methods. The results demonstrate that the entrance of Glu from blood into brain is rather limited in comparison to that of neutral amino acids (al-Sarraf et al., 1995, 1997a,b; al-Sarraf and Philip, 2003) and is slow (Oldendorf, 1971; Sershen and Lajtha, 1976). Thus, the BBB helps to protect the brain from changes in circulating plasma Glu, though there are areas of increased vulnerability such as the circumventricular organs that do not contain a BBB (median eminence, area postrema, subfornical organ, subcommissural organ, pineal gland, neurohypophysis (Gross and Weindl, 1987).

    View all citing articles on Scopus
    1

    Present address: Dept. of Gerontology, King's College London, Cornwall House, London SE1 8WA, UK.

    View full text