Reduction of GABA and glutamate transporter messenger RNAs in the severe-seizure genetically epilepsy-prone rat

The genetically epilepsy-prone rat is an animal model of inherited generalised tonic-clonic epilepsy that shows abnormal susceptibility to audiogenic seizures and a lowered threshold to a variety of seizure-inducing stimuli. Recent studies suggest a crucial role for glutamate and GABA transporters in epileptogenesis and seizure propagation. The present study examines the levels of expression of the messenger RNAs encoding the glial and neuronal glutamate transporters, GLT-1 and EAAC-1, and the neuronal GABA transporter, GAT-1, in paired male genetically epileptic-prone rats and Sprague Dawley control rats using the technique of in situ hybridization. In a parallel study, semiquantitative immunoblotting was used to assess GLT-1 and EAAC-1 protein levels in similarly paired animals. Animals were assessed for susceptibility to audiogenic seizures on six occasions, and killed seven days following the last audiogenic stimulus exposure. Rat brains were processed for in situ hybridization with radioactive 35S-labelled oligonucleotide probes (EAAC-1 and GAT-1), 35S-labelled riboprobes (GLT-1), and Fluorescein-labelled riboprobes (GLT-1 and GAT-1) or processed for immunoblotting using subtype-specific antibodies for GLT-1 and EAAC-1. Semiquantitative analyses were carried out on X-ray film autoradiograms in several brain regions for both in situ hybridization and immunoblotting studies. Reductions in GAT-1 messenger RNA were found in genetically epileptic-prone rats in all brain regions examined (-8 to -24% compared to control). Similar reductions in GLT-1 messenger RNA expression levels were seen in cortex, striatum, and CA1 (-8 to -12%) of genetically epileptic-prone rats; the largest reduction observed was in the inferior colliculus (-20%). There was a tendency for a reduced expression of EAAC-1 messenger RNA in most regions of the genetically epileptic-prone rat brain although this reached statistical significance only in the striatum (-12%). In contrast, no significant differences in GLT-1 and EAAC-1 protein between genetically epileptic-prone rats and control animals were observed in any region examined, although there was a tendency to follow the changes seen with the corresponding messenger RNAs. These results show differences in the messenger RNA expression levels of three crucial amino acid transporters. For the two glutamate transporters, GLT-1 and EAAC-1, differences in messenger RNA levels are not reflected or are only partially reflected in the expression of the corresponding proteins.     

Neuropeptides in the skin: interactions between the neuroendocrine and the skin immune systems

The interaction between components of the nervous system and multiple target cells in the cutaneous immune system has been receiving increasing attention. It has been observed that certain skin diseases such as psoriasis and atopic dermatitis have a neurogenic component. Neuropeptides released by sensory nerves that innervate the skin and often contact epidermal and dermal cells can directly modulate functions of keratinocytes, Langerhans cells (LC), mast cells, dermal microvascular endothelial cells and infiltrating immune cells. Among these neuropeptides the tachykinins substance P (SP) and neurokinin A (NKA), calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP) and somatostatin (SOM) have been reported to effectively modulate skin and immune cell functions such as cell proliferation, cytokine production or antigen presentation under physiological or pathophysiological conditions. Expression and regulation of their corresponding receptors that are expressed on a variety of skin cells as well as the presence of neuropeptide-specific peptidases such as neutral endopeptidase (NEP) or angiotensin-converting enzyme (ACE) determine the final biological response mediated by these peptides on the target cell or tissue. Likewise, skin cells like keratinocytes or fibroblasts are a source for neurotrophins such as nerve growth factor that are required not only for survival and regeneration of sensory neurons but also to control responsiveness of these neurons to external stimuli. Therefore, neuropeptides, neuropeptide receptors, neuropeptide-degrading enzymes and neurotrophins participate in a complex, interdependent network of mediators that modulate skin inflammation, wound healing and the skin immune system. This review will focus on recent studies demonstrating the role of tachykinins, CGRP, SOM and VIP and their receptors and neuropeptide-degrading enzymes in mediating neurogenic inflammation in the skin.     

Field aflatoxin contamination of corn in South Georgia

The weather conditions in 1977 were generally not favorable for corn production in many of the southeastern states. A combination of stress factors, weather conditions, and insect damage was favorable for ear mold by species of theAspergillus flavus group.A. flavus was the dominant fungus in preharvest corn, and this led to extensive aflatoxin contamination.     

Constitutive achlorhydria in mucolipidosis type IV

Mucolipidosis type IV is an autosomal recessive lysosomal storage disease of unknown etiology that causes severe neurological and ophthalmological abnormalities. In an attempt to obtain insight into the nature of the metabolic abnormality in this disorder, we prospectively evaluated 15 consecutive patients, aged 2 to 23 years, over a period of 22 months. The finding of iron deficiency in some of the patients led us to the discovery that all patients but one had markedly elevated blood gastrin levels. None had vitamin B12 deficiency. Gastroscopy in three patients showed normal gross appearance of the mucosa in two patients, 4 and 7 years old, and mucosal atrophy in a 22-year-old. Parietal cells were present in normal numbers and contained large cytoplasmic inclusions that were confirmed immunohistochemically to be lysosomal in nature. Other gastric epithelial cells appeared normal. Parietal cells contained very few tubulovesicular membranes, suggesting cellular activation, whereas apical canaliculi appeared relatively nonactivated. Both subunits of the parietal cell H+/K+-ATPase were present, and both partially colocalized with f-actin at the apical membrane. We conclude that patients with mucolipidosis type IV are constitutively achlorhydric and have partially activated parietal cells. We hypothesize that the defective protein in this disease is closely associated with the final stages of parietal cell activation and is critical for a specific type of cellular vacuolar trafficking between the cytoplasm and the apical membrane domain.     

Distribution of antifungal proteins in maize kernel tissues using immunochemistry

This study examined the distribution of two antifungal proteins, ribosome-inactivating protein (RIP) and zeamatin, in maize kernel tissues. Proteins were extracted from endosperm (including aleurone layer) and embryo tissues of imbibed maize kernels. Western blot analyses revealed that RIP-like protein was present at higher levels in endosperm than in embryo tissues, whereas zeamatin-like protein was more concentrated in embryo tissues than in endosperm tissues. However, there were three protein bands in the endosperm and two bands in the embryo that reacted to anti-RIP antibody in Western blot analyses. Tissue prints were conducted to localize the antifungal proteins. Imbibed kernels were cut longitudinally and transversely and blotted onto nitrocellulose membranes. Using antibodies against maize RIP and zeamatin, RIP was found primarily in the aleurone layer of the endosperm and glandular layer of scutellum, whereas zeamatin was located mainly in the kernel embryo. These results provide insight into the potential functions of these antifungal proteins, especially since the presence of RIP and zeamatin within maize kernels uniquely protects kernels from pathogens.