Mild closed head injury promotes a selective trigeminal hypernociception: implications for the acute emergence of post-traumatic headache.

BACKGROUND: Headache is one of the most common symptoms following traumatic head injury. The mechanisms underlying the emergence of such post-traumatic headache (PTH) remain unknown but may be related to injury of deep cranial tissues or damage to central pain processing pathways, as a result of brain injury. METHODS: A mild closed head injury in mice combined with the administration of cranial or hindpaw formalin tests was used to examine post-traumatic changes in the nociceptive processing from deep cranial tissues or the hindpaw. Histological analysis was used to examine post-traumatic pro-inflammatory changes in the calvarial periosteum, a deep cranial tissue. RESULTS: At 48 h after head injury, mice demonstrated enhanced nociceptive responses following injection of formalin into the calvarial periosteum, a deep cranial tissue, but no facilitation of the nociceptive responses following injection of formalin into an extracranial tissue, the hindpaw. Mice also showed an increase in the number of activated periosteal mast cells 48 h following mild head trauma, suggesting an inflammatory response. CONCLUSION: Our study demonstrates that mild closed head injury is associated with enhanced processing of nociceptive information emanating from trigeminal-innervated deep cranial tissues, but not from non-cranial tissues. Based on these finding as well as the demonstration of head injury-evoked degranulation of calvarial periosteal mast cells, we propose that inflammatory-evoked enhancement of peripheral cranial nociception, rather than changes in supraspinal pain mechanisms play a role in the initial emergence of PTH. Peripheral targeting of nociceptors that innervate the calvaria may be used to ameliorate PTH pain.

Sumatriptan alleviates nitroglycerin-induced mechanical and thermal allodynia in mice.

The association between the clinical use of nitroglycerin (NTG) and headache has led to the examination of NTG as a model trigger for migraine and related headache disorders, both in humans and laboratory animals. In this study in mice, we hypothesized that NTG could trigger behavioural and physiological responses that resemble a common manifestation of migraine in humans. We report that animals exhibit a dose-dependent and prolonged NTG-induced thermal and mechanical allodynia, starting 30-60 min after intraperitoneal injection of NTG at 5-10 mg/kg. NTG administration also induced Fos expression, an anatomical marker of neuronal activity in neurons of the trigeminal nucleus caudalis and cervical spinal cord dorsal horn, suggesting that enhanced nociceptive processing within the spinal cord contributes to the increased nociceptive behaviour. Moreover, sumatriptan, a drug with relative specificity for migraine, alleviated the NTG-induced allodynia. We also tested whether NTG reduces the threshold for cortical spreading depression (CSD), an event considered to be the physiological substrate of the migraine aura. We found that the threshold of CSD was unaffected by NTG, suggesting that NTG stimulates migraine mechanisms that are independent of the regulation of cortical excitability.

Animal models of migraine: looking at the component parts of a complex disorder.

Animal models of human disease have been extremely helpful both in advancing the understanding of brain disorders and in developing new therapeutic approaches. Models for studying headache mechanisms, particularly those directed at migraine, have been developed and exploited efficiently in the last decade, leading to better understanding of the potential mechanisms of the disorder and of the action for antimigraine treatments. Model systems employed have focused on the pain-producing cranial structures, the large vessels and dura mater, in order to provide reproducible physiological measures that could be subject to pharmacological exploration. A wide range of methods using both in vivo and in vitro approaches are now employed; these range from manipulation of the mouse genome in order to produce animals with human disease-producing mutations, through sensitive immunohistochemical methods to vascular, neurovascular and electrophysiological studies. No one model system in experimental animals can explain all the features of migraine; however, the systems available have begun to offer ways to dissect migraine's component parts to allow a better understanding of the problem and the development of new treatment strategies.

The three human syntrophin genes are expressed in diverse tissues, have distinct chromosomal locations, and each bind to dystrophin and its relatives.

The syntrophins are a biochemically heterogeneous group of 58-kDa intracellular membrane-associated dystrophin-binding proteins. We have cloned and characterized human acidic (alpha 1-) syntrophin and a second isoform of human basic (beta 2-) syntrophin. Comparison of the deduced amino acid structure of the three human isoforms of syntrophin (together with the previously reported human beta 1-syntrophin) demonstrates their overall similarity. The deduced amino acid sequences of human alpha 1- and beta 2-syntrophin are nearly identical to their homologues in mouse, suggesting a strong functional conservation among the individual isoforms, Much like beta 1-syntrophin, human beta 2-syntrophin has multiple transcript classes and is expressed widely, although in a distinct pattern of relative abundance. In contrast, human alpha 1-syntrophin is most abundant in heart and skeletal muscle, and less so in other tissues. Somatic cell hybrids and fluorescent in situ hybridization were both used to determine their chromosomal locations: beta 2-syntrophin to chromosome 16q22-23 and alpha 1-syntrophin to chromosome 20q11.2. Finally, we used in vitro translated proteins in an immunoprecipitation assay to show that, like beta 1-syntrophin, both beta 2- and alpha 1-syntrophin interact with peptides encoding the syntrophin-binding region of dystrophin, utrophin/dystrophin related protein, and the Torpedo 87K protein.

Syntrophin binds to an alternatively spliced exon of dystrophin.

Dystrophin, the protein product of the Duchenne muscular dystrophy locus, is a protein of the membrane cytoskeleton that associates with a complex of integral and membrane-associated proteins. Of these, the 58-kD intracellular membrane-associated protein, syntrophin, was recently shown to consist of a family of three related but distinct genes. We expressed the cDNA of human beta 1-syntrophin and the COOH terminus of human dystrophin in reticulocyte lysates using an in vitro transcription/translation system. Using antibodies to dystrophin we immunoprecipitated these two interacting proteins in a variety of salt and detergent conditions. We demonstrate that the 53 amino acids encoded on exon 74 of dystrophin, an alternatively spliced exon, are necessary and sufficient for interaction with translated beta 1-syntrophin in our assay. On the basis of its alternative splicing, dystrophin may thus be present in two functionally distinct populations. In this recombinant expression system, the dystrophin relatives, human dystrophin related protein (DRP or utrophin) and the 87K postsynaptic protein from Torpedo electric organ, also bind to translated beta 1-syntrophin. We have found a COOH-terminal 37-kD fragment of beta 1-syntrophin sufficient to interact with translated dystrophin and its homologues, suggesting that the dystrophin binding site on beta 1-syntrophin occurs on a region that is conserved among the three syntrophin homologues.

The cloning of zebrin II reveals its identity with aldolase C.

The sagittal organization of the mammalian cerebellum can be observed at the anatomical, physiological and biochemical level. Previous screening of monoclonal antibodies produced in our laboratory has identified two intracellular antigens, zebrin I and II, that occur exclusively in adult cerebellar Purkinje cells. As their name suggests, the zebrin antibody staining of the Purkinje cell population is not uniform. Rather, zebrin-positive Purkinje cells are organized in stripes or bands that run from anterior to posterior across most of the cerebellum; interposed between the zebrin-positive cells are bands of Purkinje cells that are zebrin-negative. Comparison of the position of the antigenic bands with the anatomy of afferent projections suggests that the bands are congruent with the basic developmental and functional 'compartments' of the cerebellum. We report the isolation of cDNA clones of the 36 x 10(3) M(r) antigen, zebrin II, by screening of a mouse cerebellum cDNA expression library. Sequence analysis reveals a 98% identity between our clone and the glycolytic isozyme, aldolase C. In order to more rigorously demonstrate the identity of the two proteins, we stained adult cerebellum with an independent monoclonal antibody raised against aldolase C. Anti-aldolase staining occurs in a previously unreported pattern of sagittal bands of Purkinje cells; the pattern is identical to that revealed by the zebrin II monoclonal. Further, in situ hybridization of antisense aldolase C riboprobe shows that the accumulation of zebrin II/aldolase C mRNA corresponds to the pattern of the zebrin antigen in Purkinje cells. Zebrin II/aldolase C gene expression is thus regulated at the level of transcription (or mRNA stability). In light of previous work that has demonstrated the cell-autonomous and developmentally regimented expression of zebrin II, further studies of the regulation of this gene may lead to insights about the determination of cerebellar compartmentation.

Cloning of human basic A1, a distinct 59-kDa dystrophin-associated protein encoded on chromosome 8q23-24.

Duchenne and Becker muscular dystrophies are caused by defects of dystrophin, which forms a part of the membrane cytoskeleton of specialized cells such as muscle. It has been previously shown that the dystrophin-associated protein A1 (59-kDa DAP) is actually a heterogeneous group of phosphorylated proteins consisting of an acidic (alpha-A1) and a distinct basic (beta-A1) component. Partial peptide sequence of the A1 complex purified from rabbit muscle permitted the design of oligonucleotide probes that were used to isolate a cDNA for one human isoform of A1. This cDNA encodes a basic A1 isoform that is distinct from the recently described syntrophins in Torpedo and mouse and is expressed in many tissues with at least five distinct mRNA species of 5.9, 4.8, 4.3, 3.1, and 1.5 kb. A comparison of our human cDNA sequence with the GenBank expressed sequence tag (EST) data base has identified a relative from human skeletal muscle, EST25263, which is probably a human homologue of the published mouse syntrophin 2. We have mapped the human basic component of A1 and EST25263 genes to chromosomes 8q23-24 and 16, respectively.

The structural and functional diversity of dystrophin.

Duchenne and Becker muscular dystrophies are caused by defects of the dystrophin gene. Expression of this large X-linked gene is under elaborate transcriptional and splicing control. At least five independent promoters specify the transcription of their respective alternative first exons in a cell-specific and developmentally controlled manner. Three promoters express full-length dystrophin, while two promoters near the C terminus express the last domains in a mutually exclusive manner. Six exons of the C terminus are alternatively spliced, giving rise to several alternative forms. Genetic, biochemical and anatomical studies of dystrophin suggest that a number of distinct functions are subserved by its great structural diversity. Extensive studies of dystrophin may lead to an understanding of the cause and perhaps a rational treatment for muscular dystrophy.

Clathrin light chains LCA and LCB are similar, polymorphic, and share repeated heptad motifs.

The clathrin light chains fall into two major classes, LCA and LCB. In an intact clathrin triskelion, one light chain, of either class, is bound to the proximal segment of a heavy chain leg. Analysis of rat brain and liver complementary DNA clones for LCA and LCB shows that the two light chain classes are closely related. There appear to be several members of each class having deletions of varying length aligned at the same position. A set of ten heptad elements, characteristic of alpha-helical coiled coils, is a striking feature of the central part of each derived amino acid sequence. These observations suggest a model in which the alpha-helical segment mediates binding to clathrin heavy chains and the amino- and carboxyl-terminal segments mediate interactions with other proteins. They also suggest an explanation for the observed tissue-dependent size variation for members of each class.