Syntaxin 1 binds to several proteins of the synaptic terminal and is a central component in the pathway of protein-protein interactions that underlies docking and fusion of synaptic vesicles. Molecular studies revealed the occurrence of two isoforms, syntaxin 1A and syntaxin 1B, which coexpress in neural tissues. However, they display differential expression patterns in endocrine cell types. We generated isoform-specific antibodies that were used in Western blotting and immunocytochemical studies. First, we confirmed the sole presence of syntaxin 1A in endocrine pituitary cells. Second, we found distinctive immunolabelling patterns of each isoform in the rat olfactory system, hippocampus, striatum, thalamus and spinal cord. In addition, the principal white matter commissures displayed distinct immunoreactivity for each isoform. This report shows, for the first time, major differences between the distributions of syntaxin 1A and syntaxin 1B isoforms in the rat central nervous system.

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System A for neutral amino acid transport is Increased by hypertonic shock in NBL-1 cells previously induced to express system A activity by amino acid starvation. The hypertonicity-medated effect can be blocked by cyclobeximide but is insensitive to tuncmycin. The activity induced may be inactivated Irreversibly by the addition of system A substrates, by a rapid mehanism insensitive to cycloheximide. In CHO-Ki cells, hypertonicity increases system A activity, as has been shown in NBL-1 cells. This effect is additive to the activity produced by derepression of system A by amino acid starvation and is insensitive to tuncmycin. Furthermore, the alanine-resistant mutant CHO-K1 alar4, which bears a mutation affecting the regulatory gene RI, involved in the derepression ofsystem A activity after amino acid starvation, is still able to respond to the hypertonic shock by increasing system A activiy to a level slmllar to that described in hypertonicityinduced derepressed CHO-Ki (wild type) cells. These results suggest (i) that the hypertonicity-medated increase of system A activity occurs o h a mechanism other than that involved in system A derepresslon and (is) that a regulatory protein coded by an osmotically sensitive gene is responsible for further activation of preexisting A carriers.

Electrophysiological, morphological, and biochemical approaches were combined to study the effect of the presynaptic injection of the light chain of botulinum toxin C1 into the squid giant synapse. Presynaptic injection was accompanied by synaptic block that occurred progressively as the toxin filled the presynaptic terminal. Neither the presynaptic action potential nor the Ca 2؉ currents in the presynaptic terminal were affected by the toxin. Biochemical analysis of syntaxin moiety in squid indicates that the light chain of botulinum toxin C1 lyses syntaxin in vitro, suggesting that this was the mechanism responsible for synaptic block. Ultrastructure of the injected synapses demonstrates an enormous increase in the number of presynaptic vesicles, suggesting that the release rather than the docking of vesicles is affected by biochemical lysing of the syntaxin molecule.Calcium-triggered fusion of synaptic vesicles with the presynaptic membrane is one of the most specialized features of neurons and is an essential step for chemical neurotransmission. Molecular mechanisms underlying this process have recently been described (1). The formation of the complex composed of the plasma membrane proteins syntaxin 1, synaptosomal-associated protein of 25 kDa (SNAP-25), and vesicle-associated membrane protein synaptobrevin constitutes a key step in the pathway to exocytosis (2). Indeed, experimental evidence for an essential role of this complex came from the identification of these proteins as substrates of neurotransmission-blocking clostridium neurotoxins (3). Furthermore, cleavage of syntaxin 1 by botulinum toxin C1 (BoNT͞C1) impairs synaptic vesicle exocytosis and blocks neurotransmitter release (4). Syntaxin 1 is present at the presynaptic active zone and is mostly associated with calcium channel proteins, in particular N (5, 6) and P͞Q (7, 8) type calcium channels, and with a possible calcium sensor, synaptotagmin, in a calciumdependent manner. Molecular studies on syntaxin 1 revealed two isoforms of this protein in mammalian neuronal tissues, syntaxin 1A and syntaxin 1B, and both isoforms are cleaved by BoNT͞C1 (3, 9). The squid Loligo pelaii expresses a presynaptic protein similar to mammalian syntaxin 1 (10). The protein was isolated from squid optic lobes, cloned, and sequenced, and a polyclonal anti-Loligo syntaxin antibody was generated that demonstrated positive immunoblot reaction against the squid syntaxin (10). Presynaptic injection of antiLoligo syntaxin IgG into the squid giant synapse results in transmitter release block (10). The dynamics of this block were of the type expected with vesicular fusion impairment (11), rather than with a reduction of vesicular availability (12, 13). In the present study, we injected presynaptically the light chain of BoNT͞C1 (BoNT͞C1-LC), known to lyse the mammalian syntaxins 1A and 1B. Presynaptic BoNT͞C1-LC injections were combined with electrophysiological, neurochemical, and ultrastructural studies to determine the effect of such toxin on transmitter release a...