3-aminocyclohexanecarboxylic-acid and 2-4-diaminobutyric-acid

Glial fibrillary acidic protein (GFAP)-positive astrocytes in preconfluent cultures derived from postnatal rat cerebellum have been previously shown to display two distinct morphologies, one stellate and the other irregularly epithelioid. The immunofluorescence studies described here showed that these cells also possess unique surface characteristics. In cultures derived from 8-day-old animals stellate cells bound the monoclonal antibody A2B5 whereas the epithelioid cells bound another monoclonal antibody against rat neural antigen-2 (RAN2). Some stellate cells derived from 2-day-old animals also bound tetanus toxin. The A2B5 labelling of the stellate cells made it possible to follow their fate in vitro. In confirmation of previous time-lapse studies, they underwent a shape transformation as confluence was approached, ultimately attaining a form resembling that of the epithelioid cells. Autoradiographic transport studies using two tritiated gamma-aminobutyric acid (GABA) analogues cis-1,3-aminocyclohexane carboxylic acid (ACHC) and beta-alanine revealed further differences between the two types of astrocytes. Whereas [3H]ACHC was taken up solely by the stellate cells [3H]beta-alanine was transported by both cell types. In other experiments in which various inhibitors of [3H]GABA transport were used ACHC virtually eliminated uptake into the stellate astrocyte, but had little effect on the epithelioid ones. The 'neuron-like' [3H]GABA transport process in the stellate astrocytes was confirmed in experiments comparing the effect of another compound which has been proposed as an astrocyte-selective GABA transport inhibitor, 4,5,6,7-tetrahydroisoxazolo-(4,5-C)pyridin-3-ol (THPO). No discrimination was found in its effect on the uptake of [3H]GABA into either neurons or stellate astrocytes. Further autoradiographic studies following the uptake of [3H]GABA by postnatal cerebellar slices showed that astrocytes in all layers of the cerebellar cortex and white matter transported [3H]GABA in contrast to the situation in culture where the amino acid is taken up predominantly by the stellate astrocytes. The possibility is discussed that the stellate astrocytes represent a population of cerebellar fibrous astrocytes whereas the identity of the epithelioid astrocytes is less certain.

Topics: Amino Acids; Amino Acids, Cyclic; Aminobutyrates; Animals; Antigens, Surface; Astrocytes; Autoradiography; Biological Transport, Active; Cells, Cultured; Cerebellum; Cyclohexanecarboxylic Acids; gamma-Aminobutyric Acid; Glial Fibrillary Acidic Protein; Rats; Staining and Labeling; Tetanus Toxoid

High affinity uptake, and the distribution of 3H-radiolabelled gamma-aminobutyrate (GABA), cis-3-aminocyclohexanecarboxylic acid, beta-alanine, proline, and leucine have been examined autoradiographically in laminar preparations of the myenteric plexus from the guinea-pig intestine. Following labelling with [3H]proline and [3H]leucine, which are incorporated into neurons, silver grains were concentrated over recognisable perikarya in the ganglia and meshworks of the plexus, whilst [3H]GABA labelled a smaller proportion of neurons and their processes. Specificity of labelling in the sites of [3H]GABA-uptake was established using combinations of labelled and unlabelled GABA, beta-alanine, and cis-3-aminocyclohexanecarboxylic acid, substrates for glial or neuronal high affinity GABA uptake systems. Only myenteric neurons and their processes were labelled significantly by [3H]GABA and its analogue cis-3-[3H]aminocyclohexanecarboxylic acid. Using autoradiographs of laminar preparations and paraffin sections, [3H]GABA labelling was found over nerve fibre bundles that could be traced from their ganglionic origins through the interconnecting meshworks of the myenteric plexus into the innervation of the deep muscular plexus of the circular muscle layer where GABA is evidently concerned with prejunctional modulation of transmitter release. The extensive but selective distribution of [3H]GABA high affinity uptake sites in neural elements of the guinea-pig myenteric plexus is consistent with GABA being an enteric neurotransmitter.

Topics: Alanine; Amino Acids; Amino Acids, Cyclic; Aminobutyrates; Animals; Autoradiography; Colon; Cyclohexanecarboxylic Acids; Female; gamma-Aminobutyric Acid; Guinea Pigs; Ileum; Leucine; Male; Myenteric Plexus; Neurons; Nipecotic Acids; Paraffin; Proline

Topics: Amino Acids; Amino Acids, Cyclic; Aminobutyrates; Animals; Anura; Autoradiography; Cyclohexanecarboxylic Acids; Guinea Pigs; Rabbits; Rats; Retina; Tritium