Identification of novel regulatory partners of the glutamate transporter GLT-1.

We used proximity-dependent biotin identification (BioID) to find proteins that potentially interact with the major glial glutamate transporter, GLT-1, and we studied how these interactions might affect its activity. GTPase Rac1 was one protein identified, and interfering with its GTP/GDP cycle in mixed primary rat brain cultures affected both the clustering of GLT-1 at the astrocytic processes and the transport kinetics, increasing its uptake activity at low micromolar glutamate concentrations in a manner that was dependent on the effector kinase PAK1 and the actin cytoskeleton. Interestingly, the same manipulations had a different effect on another glial glutamate transporter, GLAST, inhibiting its activity.

The Cdk5-p35 kinase associates with the Golgi apparatus and regulates membrane traffic.

We show here that an active Cdk5-p35 kinase is present in Golgi membranes, where it associates with a detergent-insoluble fraction containing actin. In addition, Cdk5-p35-dependent phosphorylation of alpha-PAK immunoreactive protein species was detected in Golgi membranes, as well as an interaction with the small GTPase, Cdc42. Moreover, antisense oligonucleotide suppression of Cdk5 or p35 in young cultured neurons, as well as inhibition of Cdk5 activity with olomoucine, blocks the formation of membrane vesicles from the Golgi apparatus.

Long-term rat survival after malignant brain tumor regression by retroviral gene therapy.

Total regression of malignant brain tumors was observed in Wistar rats after retrovirus-mediated gene therapy. Tumors were induced by inoculation of C6 rat glioblastoma cells to a specific location in the rat brain and the tumors that developed were visualized by magnetic resonance imaging (MR). Retroviral vectors were constructed from a defective murine retrovirus to which the thymidine kinase (tk 1) gene from herpes simplex was added (HSV1tk).

Role of phosphorylated MAPlB in neuritogenesis.

The distribution of microtubule-associated protein lB (MAPlB) phosphorylated by either proline-directed protein kinase (PDPK) or casein kinase II (CK II) in neuroblastoma cells and hippocampal neurons has been studied by immunofluorescence using specific antibodies to distinct phosphorylation-sensitive epitopes. A proximo-distal gradient of increasing PDPK-catalyzed phosphorylation of MAPlB is superimposed on a proximo distal gradient of decreasing CK II-catalyzed MAPlB phosphorylation within growing axon-like neurites. Additionally, CK II-phosphorylated MAPlB is present in cell bodies and dendrites where no PDPK-phosphorylated MAPlB is observed.

Effect of bilirubin on the membrane potential of rat brain synaptosomes.

The effect of the neurotoxic pigment bilirubin on the membrane potential of rat brain synaptosomes was studied by using the tetraphenylphosphonium ion (TTP+) technique. Bilirubin induces a rapid depolarization of synaptosomes, as reflected by an efflux of previously accumulated [3H]TTP+. This phenomenon persisted when the membrane potential across either the plasma membrane of the synaptosome or the inner membrane of the entrapped mitochondria was selectively depressed, thus indicating that both components of the synaptosomal membrane potential were affected by bilirubin.

Bovine chromaffin granules: immunological studies with antisera against neuropeptide Y, [Met]enkephalin and bombesin.

Antisera against neuropeptide Y, [Met]enkephalin and bombesin were used for characterizing the immunoreactive material in subcellular fractions of bovine adrenal medulla. Neuropeptide Y was identified by high performance liquid chromatography and by immunoblotting. Subcellular fractionation established that neuropeptide Y is present in chromaffin granules.

Developmental studies on the uptake of tyrosine by synaptosomes and plasma membrane vesicles derived from rat brain. Effect of thyroid hormones.

The uptake of L-tyrosine at various stages of development was examined in synaptosomes and in plasma membrane vesicles derived from rat brain. The total uptake has two components, Na+-dependent and Na+-independent, respectively. The Na+-dependent component of the transport system appears around the 5th postnatal day and increases with age.

Ontogenetic studies on tryptophan transport into plasma membrane vesicles derived from rat brain synaptosomes: effect of thyroid hormones.

The uptake of tryptophan at various stages of development was examined in plasma membrane vesicles derived from rat brain. The total uptake has two components Na+-dependent and Na+-independent respectively. The Na+-dependent component of the transport system appears around the 5th postnatal day and increases with the age.

Effect of postnatal anoxia on bilirubin levels in rat brain.

The effects of a period of anoxia 18-24 h after birth on bilirubin levels in rat brain were investigated during anoxia, recovery, and development. Postnatal anoxia induces a significant, temporary increase (up to 200% with respect to control values) in newborn rat brain bilirubin levels during anoxia and short-term recovery. Pretreatment of the newborn rats with a single dose of the drug sulfixosazole markedly enhances bilirubin accumulation in the brain of the anoxic rats.

Effect of thyroid hormones on the malic enzyme activity in rat brain during development.

Changes in the development pattern of cytoplasmic and mitochondrial malate dehydrogenase (decarboxylating; NADP+) activity in the brain of hypothyroid rats and the effect of triiodothyronine on the enzyme activity have been investigated. Hypothyroid rats showed lower malate dehydrogenase activity than controls during the suckling period. Results suggest that thyroid hormones promote the development of malic enzyme in the brain.

Effect of thyroid hormones on the 3-oxo acid CoA-transferase activity in rat brain during development.

The changes in the developmental pattern of 3-oxo acid CoA-transferase activity in the brain of hypothyroid rats and the effect of triiodothyronine on this enzyme activity have been investigated. Hypothyroid rats showed lower activity than controls during the suckling period. However, higher enzyme levels were found in treated rats after weaning in contrast to control animals.