bluuebunny/crossref_metadata_2025 huggingface dataset:
Aggregate formation in the spinal cord of mutant SOD1 transgenic mice is reversible and mediated by proteasomes
Authors: Puttaparthi Krishna , Wojcik Cezary , Rajendran Bhagya , DeMartino George N. , Elliott Jeffrey L.
Published: November 2003

Cu,Zn superoxide dismutase (SOD1) mutations cause one form of familial amyotrophic lateral sclerosis by a toxic gain of function that may be related to abnormal protein folding and aggregate formation. However, the processing pathways involved in SOD1 aggregate generation within spinal cord remain unclear. We have now developed an experimental system for studying SOD1 aggregate formation and clearance in intact spinal cord tissue. Here we demonstrate that the formation of SOD1‐positive aggregates in G93A SOD1 transgenic mouse spinal cord tissue involves proteasome‐mediated proteolysis. Organotypic spinal cord slices from 9‐day‐old transgenic mice expressing G93A SOD1 develop SOD1 aggregates with proteasome inhibition. In contrast, SOD1 aggregates do not form in spinal cord slices from wild type mice or transgenic mice overexpressing wild type SOD1 following proteasome inhibition. Furthermore, SOD1 aggregate formation within G93A SOD1 spinal cord is both sensitive to small changes in overall proteasome activity and reversible with the restoration of proteasome function. Our results also establish that adult mouse spinal cord exhibits a relative deficiency in proteasome activity compared with non‐CNS tissue that may help explain the propensity of spinal cord to form SOD1‐positive aggregates.
Activated Notch1 associates with a presenilin‐1/γ‐secretase docking site
Authors: Ramdya Pavan , Skoch Jesse , Bacskai Brian J. , Hyman Bradley T. , Berezovska Oksana
Published: November 2003

Presenilin‐1 (PS1), implicated as the active component of the γ‐secretase enzymatic complex, is known to cleave the cell surface receptor Notch1 after ligand binding. Here we directly visualize Notch1–PS1 interactions using a novel fluorescence lifetime imaging microscopy assay to monitor fluorescence resonance energy transfer. We demonstrate that endogenous Notch1 and PS1 move into close proximity at the cell surface after activation of Notch1 by the Delta1 ligand. A constitutively active N‐terminally truncated form of Notch1, an immediate substrate of the γ‐secretase complex, similarly is found in close proximity to PS1. Interestingly, this interaction remains in the presence of a potent γ‐secretase active site inhibitor. Thus ligand binding to Notch1 appears to result in access of truncated Notch1 to a putative docking site on the PS1–γ‐secretase complex. These results suggest a novel mechanism of ligand binding‐mediated signal transduction of Notch1.
Cleavage of recombinant proenkephalin and blockade mutants by prohormone convertases 1 and 2: an in vitro specificity study
Authors: Peinado Juan R. , Li Hong , Johanning Karla , Lindberg Iris
Published: November 2003

Proenkephalin (PE) derived‐peptides are thought to be generated predominantly through endoproteolytic cleavage by prohormone convertases 1 and 2 (PC1 and PC2). In order to compare cleavage site preferences of these convertases, we studied the processing of recombinant wild‐type rat PE and of two mutant PEs by recombinant purified mouse PC1 and PC2. Western blot analyses of timed digestions showed that both mouse PC1 and PC2 were able to produce a variety of large and intermediate sized‐peptides from wild‐type PE as well as from the precursors mutated at initial blockade sites. PC2 exhibited a broader specificity against PE than PC1, generating a much greater number of peptide products. Mass spectrometric identification of cleavage products showed that PC2 appeared to be the principal enzyme involved in the generation of smaller active opioids. Both enzymes were able to cleave various KR‐ and KK‐containing sites, but PC2 was also able to cleave efficiently at an RR‐V site and a KK‐M site not cleaved by PC1, suggesting the exclusion of large aliphatic residues at the P1′ position in PC1 cleavage. Alternative cleavage sites were readily chosen by convertases in blockade mutants, confirming in vivo results that cleavages do not follow an obligatory order. Furthermore, glycosylated PE was less efficiently processed by PC2, indicating that glycosylation may serve as a mechanism to hinder processing.
Thel‐isomer‐selective transport of aspartic acid is mediated by ASCT2 at the blood–brain barrier
Authors: Tetsuka Kazuhiro , Takanaga Hitomi , Ohtsuki Sumio , Hosoya Ken‐ichi , Terasaki Tetsuya
Published: November 2003

Aspartic acid (Asp) undergoesl‐isomer‐selective efflux transport across the blood–brain barrier (BBB). This transport system appears to play an important role in regulatingl‐ andd‐Asp levels in the brain. The purpose of this study was to identify the responsible transporters and elucidate the mechanism forl‐isomer‐selective Asp transport at the BBB. Thel‐isomer‐selective uptake of Asp by conditionally immortalized mouse brain capillary endothelial cells used as anin vitromodel of the BBB took place in an Na+‐ and pH‐dependent manner. This process was inhibited by system ASC substrates such asl‐alanine andl‐serine, suggesting that system ASC transporters, ASCT1 and ASCT2, are involved in thel‐isomer selective transport. Indeed,l‐Asp uptake by oocytes injected with either ASCT1 or ASCT2 cRNA took place in a similar manner to that in cultured BBB cells, whereas no significantd‐Asp uptake occurred. Although both ASCT1 and ASCT2 mRNA were expressed in the cultured BBB cells, the expression of ASCT2 mRNA was 6.7‐fold greater than that of ASCT1. Moreover, immunohistochemical analysis suggests that ASCT2 is localized at the abluminal side of the mouse BBB. These results suggest that ASCT2 plays a key role inl‐isomer‐selective Asp efflux transport at the BBB.
Immunotherapy as a therapeutic treatment for neurodegenerative disorders
Authors: White Anthony R. , Hawke Simon H.
Published: November 2003

Human neurodegenerative illnesses such as Alzheimer's disease and Creutzfeldt‐Jakob disease exact an enormous cost on individuals, families and society. For these and related disorders, current treatment is largely symptomatic without influencing the underlying disease process. Until recently, the development of immunotherapeutic approaches to neurodegenerative disorders had been almost completely ignored despite growing successes against other non‐infectious diseases such as cancer. However, since Schenk and colleagues described the antibody‐mediated clearance of amyloid plaques in a transgenic mouse model of Alzheimer's disease, a number of studies have confirmed the feasibility of this strategy for several neurodegenerative disorders including Huntington's disease and prion diseases. These reports offer the exciting prospect that either the immune system or its derivative components can be harnessed to fight the misfolded and/or aggregated proteins that accumulate in many neurodegenerative illnesses. If the remarkable power of clonal expansion, specificity and efficiency of the immune system can successfully inactivate these abnormal proteins, real hope exists that effective immunotherapeutic treatments for neurodegenerative illnesses may be available in the near future.
Dopamine D1 receptors mediate CREB phosphorylation via phosphorylation of the NMDA receptor at Ser897–NR1
Authors: Dudman Joshua T. , Eaton Molly E. , Rajadhyaksha Anjali , Macías Wendy , Taher Muffadal , Barczak Amy , Kameyama Kimihiko , Huganir Richard , Konradi Christine
Published: November 2003

Addictive drugs such as amphetamine and cocaine stimulate the dopaminergic system, activate dopamine receptors and induce gene expression throughout the striatum. The signal transduction pathway leading from dopamine receptor stimulation at the synapse to gene expression in the nucleus has not been fully elucidated. Here, we present evidence that D1 receptor stimulation leads to phosphorylation of the transcription factor Ca2+ and cyclic AMP response element binding protein (CREB) in the nucleus by means of NMDA receptor‐mediated Ca2+ signaling. Stimulation of D1 receptors induces the phosphorylation of Ser897 on the NR1 subunit by protein kinase A (PKA). This phosphorylation event is crucial for D1 receptor‐mediated CREB phosphorylation. Dopamine cannot induce CRE‐mediated gene expression in neurons transfected with a phosphorylation‐deficient NR1 construct. Moreover, stimulation of D1 receptors or increase in cyclic AMP levels leads to an increase in cytosolic Ca2+ in the presence of glutamate, but not in the absence of glutamate, indicating the ability of dopamine and cyclic AMP to facilitate NMDA channel activity. The recruitment of the NMDA receptor signal transduction pathway by D1 receptors may provide a general mechanism for gene regulation that is fundamental for mechanisms of drug addiction and long‐term memory.
The pesticide rotenone induces caspase‐3‐mediated apoptosis in ventral mesencephalic dopaminergic neurons
Authors: Ahmadi Ferogh A. , Linseman Daniel A. , Grammatopoulos Tom N. , Jones Susan M. , Bouchard Ron J. , Freed Curt R. , Heidenreich Kim A. , Zawada W. Michael
Published: November 2003

In vivo, the pesticide rotenone induces degeneration of dopamine neurons and parkinsonian‐like pathology in adult rats. In the current study, we utilized primary ventral mesencephalic (VM) cultures from E15 rats as an in vitro model to examine the mechanism underlying rotenone‐induced death of dopamine neurons. After 11 h of exposure to 30 nm rotenone, the number of dopamine neurons identified by tyrosine hydroxylase (TH) immunostaining declined rapidly with only 23% of the neurons surviving. By contrast, 73% of total cells survived rotenone treatment, indicating that TH+ neurons are more sensitive to rotenone. Examination of the role of apoptosis in TH+ neuron death, revealed that 10 and 30 nm rotenone significantly increased the number of apoptotic TH+ neurons from 7% under control conditions to 38 and 55%, respectively. The increase in apoptotic TH+ neurons correlated with an increase in immunoreactivity for active caspase‐3 in TH+ neurons. The caspase‐3 inhibitor, DEVD, rescued a significant number of TH+ neurons from rotenone‐induced death. Furthermore, this protective effect lasted for at least 32 h post‐rotenone and DEVD exposure, indicating lasting neuroprotection achieved with an intervention prior to the death commitment point. Our results show for the first time in primary dopamine neurons that, at low nanomolar concentrations, rotenone induces caspase‐3‐mediated apoptosis. Understanding the mechanism of rotenone‐induced apoptosis in dopamine neurons may contribute to the development of new neuroprotective strategies against Parkinson's disease.
Diazoxide induces delayed pre‐conditioning in cultured rat cortical neurons
Authors: Kis Bela , Rajapakse Nishadi C. , Snipes James A. , Nagy Krisztina , Horiguchi Takashi , Busija David W.
Published: November 2003

We investigated the effect of diazoxide on neuronal survival in primary cultures of rat cortical neurons against oxygen–glucose deprivation (OGD). Diazoxide pre‐treatment induced delayed pre‐conditioning and almost entirely attenuated the OGD‐induced neuronal death. Diazoxide inhibited succinate dehydrogenase and induced mitochondrial depolarization, free radical production and protein kinase C activation. The putative mitochondrial ATP‐sensitive potassium channel blocker 5‐hydroxydecanoate abolished the protective effect of diazoxide while the non‐selective KATP channel blocker glibenclamide did not. The non‐selective KATP channel openers nicorandil and cromakalim did not improve viability. Superoxide dismutase mimetic, M40401, or protein kinase C inhibitor, chelerythrine, prevented the neuroprotective effect of diazoxide. Diazoxide did not increase reduced glutathione and manganese‐superoxide dismutase levels but we found significantly higher reduced glutathione levels in diazoxide‐pre‐conditioned neurons after OGD. In pre‐conditioned neurons free radical production was reduced upon glutamate stimulation. The succinate dehydrogenase inhibitor 3‐nitropropionic acid also induced pre‐conditioning and free radical production in neurons. Here, we provide the first evidence that diazoxide induces delayed pre‐conditioning in neurons via acute generation of superoxide anion and activation of protein kinases and subsequent attenuation of oxidant stress following OGD. The succinate dehydrogenase‐inhibiting effect of diazoxide is more likely to be involved in this neuroprotection than the opening of mitochondrial ATP‐sensitive potassium channels.
The myelin–axolemmal complex: biochemical dissection and the role of galactosphingolipids
Authors: Menon Krishna , Rasband Matthew N. , Taylor Christopher M. , Brophy Peter , Bansal Rashmi , Pfeiffer Steven E.
Published: November 2003

Myelin–axolemmal interactions regulate many cellular and molecular events, including gene expression, oligodendrocyte survival and ion channel clustering. Here we report the biochemical fractionation and enrichment of distinct subcellular domains from myelinated nerve fibers. Using antibodies against proteins found in compact myelin, non‐compact myelin and axolemma, we show that a rigorous procedure designed to purify myelin also results in the isolation of the myelin–axolemmal complex, a high‐affinity protein complex consisting of axonal and oligodendroglial components. Further, the isolation of distinct subcellular domains from galactolipid‐deficient mice with disrupted axoglial junctions is altered in a manner consistent with the delocalization of axolemmal proteins observed in these animals. These results suggest a paradigm for identification of proteins involved in neuroglial signaling.
Caspase activation contributes to delayed death of heat‐stressed striatal neurons
Authors: White Michael G. , Emery Michael , Nonner Doris , Barrett John N.
Published: November 2003

Hyperthermia can contribute to brain damage both during development and post‐natally. We used rat embryonic striatal neurons in culture to study mechanisms underlying hyperthermia‐induced neuronal death. Heat stress at 43°C for 2 h produced no obvious signs of damage during the first 12 h after the stress, but more than 50% of the neurons died during the next 3 days. More than 40% of the neurons had activated caspases 24 h following the heat stress. Caspase‐3 activity increased with a delay of more than 10 h following cessation of the heat stress, reaching a peak at ∼18 h. Neuronal death measured 1–3 days after the stress was reduced by the general caspase inhibitors qVD‐OPH (10–20 µm) and zVAD‐fmk (50–100 µm). These inhibitors were protective even when added 9 h after cessation of the heat stress, consistent with the delayed activation of caspases. In contrast, blockers of Na+ channels and ionotropic glutamate receptors did not reduce the heat‐induced death, indicating that glutamate excitotoxicity was not required for this neuronal death. These results show that the neuronal death produced by heat stress has characteristics of apoptosis, and that caspase inhibitors can delay this death.
Free radical‐dependent nuclear localization of Bcl‐2 in the central nervous system of aged rats is not associated with Bcl‐2‐mediated protection from apoptosis
Authors: Kaufmann Joel A. , Perez Mike , Zhang WenRu , Bickford Paula C. , Holmes Douglas B. , Taglialatela Giulio
Published: November 2003

We have previously reported that Bcl‐2 is up‐regulated in the CNS of aged F344 rats as a consequence of oxidative stress. In addition to increased levels of expression, we now report that there is a subcellular redistribution of Bcl‐2 in the CNS of aged F344 rats. Using western blotting, we found Bcl‐2 predominantly located in the cytosol of young rats. However, in aged rats Bcl‐2 was found primarily in the nucleus. This distribution, in the hippocampus and cerebellum, was reversed by treatment with the nitrone spin trap N‐tert‐butyl‐α‐phenylnitrone (PBN). Paradoxically, PBN treatment in young rats had the opposite effect, changing Bcl‐2 from predominantly cytosolic to nuclear. We also detected an increase in Bax in aged hippocampal samples (both nuclear and cytosolic), which was reversed by treatment with PBN. The distribution of Bcl‐2 and Bax in the cytosol of aged rats dramatically decreased the Bcl‐2/Bax ratio, a probable indicator of neuronal vulnerability, which was restored upon treatment with PBN. In order to assess the effect of nuclear association of Bcl‐2 we used PC12 cells stably transfected with a Bcl‐2 construct to which we added the nuclear localization sequence of the SV40 large T antigen to the N‐terminus which resulted in nuclear targeting of Bcl‐2. Measurement of cell death using lactate dehydrogenase assays showed that, contrary to wild‐type Bcl‐2, Bcl‐2 localized to the nucleus was not effective in protecting cells from treatment with 250 µm H2O2. These results suggest that nuclear localization of Bcl‐2 observed in the aged CNS may not reflect a protective mechanism against oxidative stress, a major component of age‐associated CNS impairments.
Androgens modulate β‐amyloid levels in male rat brain
Authors: Ramsden M. , Nyborg A. C. , Murphy M. P. , Chang L. , Stanczyk F. Z. , Golde T. E. , Pike C. J.
Published: November 2003

As a normal consequence of aging, men experience a significant decline in androgen levels. Although the neural consequences of age‐related androgen depletion remain unclear, recent evidence suggests a link between low androgen levels and the development of Alzheimer's disease (AD). Here, we test the hypothesis that androgens act as endogenous modulators of β‐amyloid protein (Aβ) levels. To investigate this possibility, brain and plasma levels of Aβ were measured in male rats with varying hormonal conditions. Depletion of endogenous sex steroid hormones via gonadectomy (GDX) resulted in increased brain levels of Aβ in comparison to gonadally intact male rats. This GDX‐induced increase in Aβ levels was reversed by DHT supplementation, demonstrating a functional role for androgens in modulating brain levels of Aβ. These findings suggest that age‐related androgen depletion may result in accumulation of Aβ in the male brain and thereby act as a risk factor for the development of AD.
Short Course of Prednisolone in Indian Patients with Solitary Cysticercus Granuloma and New‐onset Seizures
Authors: Mall Ravi Kant , Agarwal Atul , Garg Ravindra Kumar , Kar Alok Mohan , Shukla Rakesh
Published: November 2003

Summary:  Purpose: To evaluate the role of a short course of oral corticosteroids in Indian patients with solitary cysticercus granuloma with seizures.

Methods: In this open‐label, randomized, prospective follow‐up study, 97 patients with new‐onset seizures and a single enhancing computed tomography (CT)‐detected lesion of cysticercosis were randomly divided in two groups to receive either antiepileptic monotherapy alone (n = 48) or antiepileptic monotherapy with prednisolone (n = 49). The patients in the latter group received prednisolone, 1 mg/kg/day for 10 days, followed by tapering over next 4 days. The patients were followed up for 6 months. Repeated CT scans were performed after 1 and 6 months.

Results: The majority of patients were young. Simple partial seizure, with or without secondary generalization, was the commonest seizure type encountered. Follow‐up CT scans at 1 and 6 months demonstrated a significantly better response for prednisolone as far as complete resolution of CT lesion was concerned. Kaplan–Meier analysis suggested significantly less probability of seizure recurrence for prednisolone‐treated patients. At 6 months, Kaplan–Meier estimated risk of seizure after first seizure was 2% in prednisolone‐treated patients in comparison to 13% for those who were not given prednisolone.

Conclusions: Short‐term prednisolone therapy helps in rapid resolution of solitary cysticercus granuloma in Indian patients with new‐onset seizures. Resolution of lesions is associated with improved seizure‐related prognosis.
The Multicenter Study of Epilepsy Surgery: Recruitment and Selection for Surgery
Authors: Berg Anne T. , Vickrey Barbara G. , Langfitt John T. , Sperling Michael R. , Walczak Thaddeus S. , Shinnar Shlomo , Bazil Carl W. , Pacia Steven V. , Spencer Susan S. ,   Multicenter Study of Epilepsy Surgery
Published: November 2003

Summary:  Purpose: Multiple studies have examined predictors of seizure outcomes after epilepsy surgery. Most are single‐center series with limited sample size. Little information is available about the selection process for surgery and, in particular, the proportion of patients who ultimately have surgery and the characteristics that identify those who do versus those who do not. Such information is necessary for providing the epidemiologic and clinical context in which epilepsy surgery is currently performed in the United States and in other developed countries.

Methods: An observational cohort of 565 surgical candidates was prospectively recruited from June 1996 through January 2001 at six Northeastern and one Midwestern surgical centers. Standardized eligibility criteria and protocol for presurgical evaluations were used at all seven sites.

Results: Three hundred ninety‐six (70%) study subjects had resective surgery. Clinical factors such as a well‐localized magnetic resonance imaging (MRI) abnormality and consistently localized EEG findings were most strongly associated with having surgery. Of those who underwent intracranial monitoring (189, 34%), 85% went on to have surgery. Race/ethnicity and marital status were marginally associated with having surgery. Age, education, and employment status were not. Demographic factors had little influence over the surgical decision. More than half of the patients had intractable epilepsy for ≥10 years and five or more drugs had failed by the time they initiated their surgical evaluation. During the recruitment period, eight new antiepileptic drugs were approved by the Food and Drug Administration for use in the United States and came into increasing use in this study's surgical candidates. Despite the increased availability of new therapeutic options, the proportion that had surgery each year did not fluctuate significantly from year to year. This suggests that, in this group of patients, the new drugs did not provide a substantial therapeutic benefit.

Conclusions: Up to 30% of patients who undergo presurgical evaluations for resective epilepsy surgery ultimately do not have this form of surgery. This is a group whose needs are not currently met by available therapies and procedures. Lack of clear localizing evidence appears to be the main reason for not having surgery. To the extent that these data can address the question, they suggest that repeated attempts to control intractable epilepsy with new drugs will not result in sustained seizure control, and eligible patients will proceed to surgery eventually. This is consistent with recent arguments to consider surgery earlier rather than later in the course of epilepsy. Postsurgical follow‐up of this group will permit a detailed analysis of presurgical factors that predict the best and worst seizure outcomes.
Infantile Spasms in a Patient with Williams Syndrome and Craniosynostosis
Authors: Morimoto Masafumi , An Byongmun , Ogami Aya , Shin Noriko , Sugino Yuriko , Sawai Yasuko , Usuku Tomohiro , Tanaka Masayuki , Hirai Kiyoshi , Nishimura Akira , Hasegawa Koh , Sugimoto Tohru
Published: November 2003

Summary:  A patient with Williams syndrome, craniosynostosis, and infantile spasms is described. At age 6 months, the infant demonstrated infantile spasms and craniosynostosis and was operated on for craniosynostosis and treated with adrenocorticotropic hormone (ACTH) for the infantile spasms. ACTH completely controlled the seizures, but was halted because of the progression of ventricular hypertrophy. The seizure returned, and he was found to have elfin face, failure‐to‐thrive, developmental delay, and dental malformation in addition to congenital heart defects. High‐resolution chromosome analysis revealed interstitial deletion of 7q11.22‐q11.23. Therefore his clinical and cytogenetic diagnosis was Williams syndrome. Thyrotropin‐releasing hormone (TRH) therapy reduced his seizures and improved the findings of EEG without cardiac side effects. In addition, his psychomotor development was slightly improved.
INFLUENCE OF ANTHROPOGENIC POLLUTION ON MACROPHYTOBENTHOS OF THE AVACHA BAY (SOUTHEAST KAMCHATKA, RUSSIA)
Authors: Klotchkova N.G. , Berezovskaya V.A.
Published: November 2003

Most industrial and agricultural enterprises of the Kamchatsky district are situated along the coast of the Avacha Bay and its river drainage. Intensive water pollution resulted in extreme deterioration of the bay's ecology. The destruction of macrophytobenthos under the influence of pollution was investigated. Materials were regularly collected at the same sampling sites in different seasons of the year from 1988 up to 1999. Analysis of our data allowed us to determine directions of long‐term changes of species composition and macrophytobenthos structure, speed of degradation of different types of vegetational communities and reasons for all these processes. The Avacha Bay's flora consisted of more than 160 species up to 1970s, including species belonged to 104 genera, 47 families, 23 orders, 3 divisions (30 Chlorophyta, 48 Phaeophyta, 82 Rhodophyta). Flora of the internal part of the bay differed from that of the bay's mouth and consisted of 143 and 146 species, respectively. Presently, the whole flora of the bay (internal part+ throat) includes only 94 species. Along the urban coast 18 poly‐ and mesosaprobious species are found only. They are characterized by small sizes, low productivity and short life cycles. Most of them are ephemeral with a primitive morphology. The responsive reaction of algae to pollution resulted in decreases in size and weight of the thalli, and appearance of anomalies of development. Laminaria bongardiana was used as a bioindicator. The changes of weight and sizes of its plants allow us to determine degree of the pollution in different parts of the coast of the Avacha Bay.
NUCLEAR RIBOSOMAL DNA PHYLOGENY OF HALIMEDALES REVEALS SEVERAL POLYPHYLETIC GENERA
Authors: Kooistra W. H. C.F. , Hillis L.W.
Published: November 2003

Halimedales (Chlorophyta) are widely distributed in reefs and lagoons throughout the tropics and subtropics. One genus, Chlorodesmis, consists of simple, branching siphons that persist into the mature sexual phase. In most other genera, only the juvenile stages exhibit such simple anatomy. Their mature, often‐calcified thalli possess complex siphonal anatomy and differentiation depending on the location in the thallus. We obtained a nuclear ribosomal DNA phylogeny encompassing many genera in order to unravel the evolution of this complexity. Results pinpoint Caulerpa as outgroup to a clade with all other genera included in this study. The first dichotomy separates Halimeda from the remaining genera. Distinct gametangial anatomy defines these two principal clades. Within the remainder, Tydemania and its sister Flabellia are nearest neighbors to a clade containing Rhipocephalus, Udotea, Penicillus and Chlorodesmis. The latter three genera are polyphyletic. Therefore, character states of thallus superstructure defining these genera appear to be homoplasies. Chlorodesmis represents a case of neoteny where the juvenile stage persists into reproductive maturity. We will evaluate which morphological characters do change state in agreement with the molecular phylogeny and can, therefore, serve as diagnostic characters for a more natural classification.
PRELIMINARY EVALUATION OF THE BIOREMEDIATION POTENTIAL OF PORPHYRA: PHOTOSYNTHETIC PRODUCTION BY BLADES AND CONCHOCELIS
Authors: Kraemer G.P. , Yarish C.
Published: November 2003

Given their rapid growth and nutrient assimilation rates, Porphyra spp. are good candidates for bioremediation. The production potential of two northeast U.S. Porphyra species currently in culture (P. purpurea and P. umbilicalis) was evaluated by measuring rates of photosynthesis (as O2 evolution) of samples grown at 20° C. Gametophytes of P. umbilicalis photosynthesized at rates that were 80% higher than those of P. purpurea over 5–20° C at both sub‐saturating and saturating irradiances (37 and 289 μmol photons m−2 s−1). Porphyra umbilicalis was both more efficient at low irradiances (higher alpha) and had a higher Pmax than did P. purpurea (23.0 vs. 15.6 μmol O2 g−1 DW min−1), suggesting that P. umbilicalis is a better choice for mass culture where self‐shading may be severe. The photosynthesis‐irradiance relationship for the Conchocelis stage of P. purpurea was also examined. Tufts of filaments, grown at 10, 15, and 20° C, were assayed at growth temperatures at irradiances ranging from 0–315 μmol photons m−2 s−1. Tufts were slightly more productive at 15° than at 10° C, but only ca. 4–6% as productive as gametophytes. Maximum rates of net photosynthesis were reduced by 66–74% in tufts grown at 20° C (only about 2% of gametophytes). The Conchocelis stage, however, need not limit mariculture operations; once Conchocelis cultures are established, they can be maintained over the long‐term as ready sources of spores for net seeding.
INTER‐FAMILY USE OF MICROSATELLITE MARKERS IN LAMINARIALES: THE EXPERIENCE BETWEEN LAMINARIA DIGITATA AND LESSONIA SPP.
Authors: Martínez E.A. , Cardenas L. , Figueroa C. , Vidal R. , Billot C.
Published: November 2003

Among the co‐dominant molecular markers, microsatellite loci have a number of advantages in population genetic studies. However, the different methods to hunt these markers are expensive, time consuming, and they require sophisticated laboratory equipment. Using in one species the microsatellites primers originally described for another one saves time and reduces costs. Examples in the literature revealed that microsatellites described for Gracilaria gracilis from France have not worked for other members of the Gracilariales. The results were not very promising, at least for Gracilaria chilensis from Chile. In this study, a number of microsatellite loci described for Laminaria digitata (Laminariaceae) from France were amplified and sequenced in two Lessonia (Lessoniaceae) species from Chile. Preliminary results show a partial conservation of both, flanking and tandem repeat regions. Some polymorphism has also been detected in Lessonia spp. The higher molecular affinity (conservation of primer sites) observed in species belonging to different families of brown algae, respect to that observed among species of the same genus in the red algae, is surprising. Such a result is strikingly counterintuitive when observing the morphological disparity among the Laminariales, respect to the similarity observed in the Gracilariales. It also reminds one of an earlier discussion by Gary Saunders about “When a family is not a family”.
ALGAL BIOREMEDIATION OF THE BERKELEY PIT LAKE
Authors: Mitman G. G.
Published: November 2003

Ongoing research is unraveling the intricacies of the microbial ecology of the Berkeley Pit Lake System, with ever increasing information becoming available regarding the diversity of Algae, Protistans, Fungi and Bacteria that inhabit this mine waste site. Defining the baseline community structure has been the first step not only toward understanding the interactions of the different groups of organisms, but also toward assessing any improvement in biodiversity within the biotic community. Now that this first step has begun, some of these extremophiles, specifically algae, that have been isolated from the Berkeley Pit Lake System are being used as a potential solution for bioremediation. The specific objectives of this research are fivefold: 1) To evaluate the bioremediative potential of our four most rapidly growing species: (Chromulina freiburgensis Dofl., Chlorella ellipsoidea Gerneck, Chlorella vulgaris Beyerinck and Chlamydomonas acidophilla Negoro) in Berkeley Pit Lake System Water with the additions of NaNO3 and NaPO4 by using an experimental matrix. This matrix will be used to estimate the minimum nutrient concentrations that would be necessary to achieve the maximum growth of algae and maximum bioremediation of the Berkeley Pit Lake System. 2) To determine which combination of nutrients will stimulate growth of the best bioremediator of our four isolated species in natural Berkeley Pit Lake System waters. In other words, what nutrient combination will give the best bioremediator a competitive edge over the other species. If time permits, different species may be grown in combination to determine if there are synergistic effects (protocooperation) between/among species. 3) To determine a temperature profile for these four species in order to determine their optimal growth temperature in Berkeley Pit Lake System water. 4) To continue to isolate organisms from the Berkeley Pit Lake System and determine their bioremediative potential. 5) Monitor algal and bacterial counts from a profile of Pit Lake System waters. The results to date will be presented for this conference.
STRUCTURAL AND SEQUENCE SIGNATURES OF NUCLEAR SSU RRNA DEFINE TAXONOMIC LEVELS WITHIN THE RHODOPHYTA
Authors: Müller K. M. , Sheath R. G. , Sherwood A.R. , Madabusi L.V. , Cannone J.J. , Subashchandran S. , Lin N. , Gutell R.R.
Published: November 2003

Over 400 nuclear SSU rRNA sequences representing all orders of the Rhodophyta were aligned and analyzed using comparative sequence analysis. Numerous nucleotide positions and structural elements were found that delineated various taxonomic groups. The 1245 region (E. coli numbering) contained a loop that differed in size between two conserved helices and clearly separated the Florideophyceae [3 nt (>95% of 268 sequences)], Bangiales [13 to 14 nt (100% of 116 sequences)] and remaining Bangiophyceae including the Cryptophyta nucleomorphs [four to eight nt (100% of 32 sequences)]. In addition, members of the Thoreaceae were found to have additional helices in the 650 and 1139 region of which a corresponding structure was not present in any other red algal SSU rRNA gene sequence. Base‐pair and nucleotide signatures differentiated the Bangiales, Florideophyceae, Bangiophyceae (not including Bangiales) and Hildenbrandiales at three levels of comparison: within the Rhodophyta (>400 sequences), the Eukaryota (not including Rhodophyta;> 1300 sequences) and three kingdom (Archaea, Bacteria, 2 organelles, Eukaryota;> 7000 sequences). For example, all members of the Hildenbrandiales have a change in the base‐pair 512:539 that is a region of functional importance. Sequences from the Eukaryota, Archaea, Bacteria and two organelles have a C:G or a U:A in this position whereas the Hildenbrandiales have a C:A pair. This analysis raises the possibility of utilizing structural features of nuclear SSU rRNA and sequence signatures to support and delineate phylogenetic clades within the Rhodophyta.
PICOPHYTOPLANKTON SEASONAL CYCLE AT THE SIO PIER, LA JOLLA, CALIFORNIA
Authors: Palenik B.P.
Published: November 2003

The abundance of phycoerythrin‐containing cyanobacteria and picoeukaryotes in water samples from the Scripps Institution of Oceanography pier have been followed at least weekly for more than two years using flow cytometry. These cyanobacteria show a seasonal cycle with generally lower cell numbers during the winter, a “bloom” as water temperatures increase, and higher cell numbers during the summer. However the population abundance appears to be more variable and the magnitude of the annual change in cell number is less than reported for coastal Massachusetts by Waterbury et al. (1986). Isolates have been obtained from pier samples and genetic characterization using rpoC1 (RNA polymerase) sequence data is in progress. The PUB:PEB chromophore ratios of isolates assayed using fluorescence excitation spectra range from about 0.4 (low PUB) to 0.7 (mid‐PUB) for isolates grown under white light. The physiological and genetic characterization of isolates is being used to examine the similarities and differences of cyanobacterial populations from different coastal regimes. Similarly a picoeukaryote has been isolated that has a flow cytometric signature similar to the natural population. It appears to be a small nonmotile prasinophyte.
SEASONAL AND ZONAL VARIATION IN NITROGEN SOURCE FOR FOUR INTERTIDAL SEAWEEDS FROM NEW ZEALAND
Authors: Phillips J.C. , Hurd C. L.
Published: November 2003

Seasonal patterns in nitrogen uptake by four species of intertidal macroalgae (Stictosiphonia arbuscula, Apophlaea lyallii, Scytothamnus australis, Xiphophora gladiata) from Otago in New Zealand's south island were investigated. Field‐collected material from species occurring at different shore heights was exposed to inorganic (NO−3, NH+4) and organic (urea) nitrogen at different concentrations (5μM, 30μM), and the rate of uptake over 30min intervals for 3hr was determined. Active uptake of nitrate was shown by all species in both seasons but uptake rates were 2–5 times greater in summer compared to winter. Ammonium uptake was highest in all species regardless of season, indicating a preference for this form of nitrogen. For all species, urea uptake was negligible during winter but represented a substantial proportion of the total nitrogen taken up during summer. Comparison of species in relation to their shore position revealed differences in uptake rates for each nitrogen source. These results suggest that intertidal seaweeds growing at different shore positions employ different strategies for procuring nitrogen in an environment where nutrient supply may be limited.
COALESCENCE VERSUS COMPETITION: FIELD AND LABORATORY STUDIES OF INTRA‐ AND INTERSPECIFIC ENCOUNTERS AMONG COALESCING SEAWEEDS
Authors: Santelices B. , Correa J.A. , Aedo D. , Hormazábal M. , Flores V.
Published: November 2003

Classical ecological theory predicts that whenever growing individuals share a common and limiting resource, such as substratum in mid‐intertidal and shallow subtidal habitats, preemptive competition will occur determining species abundance and distribution patterns. However, conspecificity of several ecologically dominant Rhodophyta may coalesce when grown in laboratory cultures. The extent at which intraspecific coalescence occurs in the field and whether the process may also happens during interspecific encounters remain to be determined. If intra‐ and interspecific coalescence effectively occurs, then coexistence through coalescence rises as an alternative to competition among red‐algal dominated intertidal and shallow subtidal communities. Populations of Mazzaella laminarioides and Nothogenia fastigiata living in mid‐intertidal, semi‐exposed rocky habitats in Central Chile are being used to test the above ideas. Intra‐ and interspecific encounters occur in the field throughout the year. Coalescence does occur among conspecific partners but it has not been detected in interspecific encounters. Rather, a thick interface of compressed cells, necrotic tissues and cyanobacterial nodules is formed between the two contacting partners. In addition, observations of laboratory cultures indicate that spore germination, germling survival and differentiation of erect axes in bispecific cultures may be reduced when compared to single‐species controls. Interspecific differences in growth and differentiation rates appear as the mechanisms explaining a lack of coalescence and negative effects during interspecific contacts. On the other hand, the existence of conspecific coalescence in the field suggests this process should be considered as a real alternative to intraspecific competition among coalescing Rhodophyta.
DIATOMS AS HOSTS FOR OTHER DIATOMS: OBSERVATIONS FROM SOUTHERN CALIFORNIA
Authors: Tiffany  , Lange C.B.
Published: November 2003

Large epiphytic and epilithic diatom species hosting other diatoms were observed in several fresh‐ and brackish water sites in Southern California. The most commonly encountered hosts were species forming long filaments attached to rocks or macroalgae, Hydrosera whampoensis (Schwarz) Hendey, Melosira varians Agardh, Pleurosira laevis (Ehrenberg) Compere and Terpsinoe musica Ehrenberg. These large diatoms often had smaller diatoms attached, usually to the girdle bands and occasionally to the mucilage pads connecting the cells. For example, cells of T. musica were observed supporting growth of a diverse diatom assemblage composed of species of the genera Achnanthes, Achnanthidium, Amphora, Cocconeis and Tabularia; Synedra sp. was attached to M. varians and B. paxillifer; and Cocconeis placentula was seen on H. whampoensis. Thus, large epiphytic and epilithic diatoms seem to provide suitable sites for attachment of small epiphytic diatom species, and it appears that this phenomenon is more common than previously thought.
MARINE MACROALGAL DIVERSITY IN THE REPUBLIC OF PANAMA
Authors: Wysor B. , Kooistra W. H. C.F. , Fredericq S.
Published: November 2003

The marine flora of Panama remains poorly described to date, having been described from a few sporadic and species poor (<50 species) collections in the early half of the century and a couple of dedicated surveys in the latter half. With the exception of a few studies that have focused on particular genera or species, only a single marine floral survey has been published in the past three decades. In 1999 we collected marine algae from nearly 100 different collection sites along both coasts of Panama over a 10 month period of time. Over 1500 specimens have been curated, representing 250 Caribbean species and 117 Pacific species. This is an increase in algal diversity of approximately 100 and 50% for the Caribbean and Pacific flora, respectively, relative to Earle's compilation of 1972. We estimate that algal diversity in Panama may approach 450 species as the remaining 30% of the collection is identified and previous records are incorporated. Aspects of biogeography will be discussed in light of such extraordinary diversity over a relatively confined region.
CHARACIOCHLORIS AND CHARACIOSIPHON FORM A SISTER GROUP TO THE “DUNALIELLA” LINEAGE
Authors: Buchheim  , Buchheim J.A.
Published: November 2003

The freshwater green algal genera, Characiochloris and Characiosiphon, are distinctive taxa for which no clear consensus exists regarding phylogenetic placement. Characiochloris is an epiphytic unicell that exhibits multiple (>2), scattered contractile vacuoles throughout the cytoplasm of the vegetative cell. Although Characiochloris has been placed in the Tetrasporales by most systematists, the uncertain status of this green algal order raises questions regarding the position of Characiochloris. Characiosiphon, a coenocytic freshwater organism, has been linked to another freshwater coenocyte, Protosiphon. However, multiple chloroplasts, greater thallus size, and a strictly aquatic life history for Characiosiphon have led to comparisons with caulerpalean ulvophytes. Phylogenetic analysis of 18S and 26S rDNA data was undertaken to test hypotheses of relationship for Characiochloris and Characiosiphon. Results from molecular phylogenetic analyses strongly support a sister relationship for Characiochloris and Characiosiphon. In addition, the Characiochloris/Characiosiphon clade is resolved as a sister group to the “Dunaliella” lineage of the Chlamydomonadales that includes Ascochloris, Chlorogonium, Dunaliella, Haematococcus, Protosiphon and Spongiochloris. These results further support the hypothesis that the “Dunaliella” clade includes the majority of taxa that exhibit multiple contractile vacuoles. Moreover, these results support an interpretation of the Characiosiphon coenocyte as a modified zoosporangium.
SOME PHYLOGENETIC RELATIONSHIPS WITHIN THE OSCILLATORIALES (CYANOBACTERIA) CLADE USING 16S RDNA GENE SEQUENCE DATA
Authors: Casamatta D.A. , Vis M. L.
Published: November 2003

An approximately 1400 base pair region of the 16S rDNA gene was sequenced from taxa within the Oscillatoriales in order to assess phylogenetic relationships. Ten previously unsequenced strains were obtained from the University of Toronto Culture Collection. New sequence data were combined with previously published sequences from a wide representation of cyanobacteria including all currently available, complete Oscillatorialian taxa. Trees constructed using parsimony, distance, and maximum likelihood methods were similar in topology, although a few taxa were variable in their placement depending on the phylogenetic method employed. Newly sequenced taxa of the genera Phormidium, Oscillatoria, and Lyngbya did not form monophyletic clades based on traditional generic designations. Two Lyngbya strains (UTCC296 and 313) and Phormidium subfuscum (UTCC474) formed a well supported monophyletic clade, but the affinity of this clade with other groups was uncertain due to lack of bootstrap support. Oscillatoria sp. (UTCC393) was closely related to the previously sequenced Oscillatoria limnetica and likewise, Phormidium molle (UTCC77) and Phormidium tenue (UTCC473) were placed in a well supported clade with other Oscillatoriales. The other four taxa were variously placed in the trees and their phylogenetic positions could not be determined with certainty.
SEXUAL REPRODUCTION AND GENETIC VARIATION IN THE SUBAERIAL ALGA CEPHALEUROS VIRESCENS (ULVOPHYCEAE, CHLOROPHYTA)
Authors: Chapman R. L. , Waters D.A. , Lopez‐Bautista J. M.
Published: November 2003

Cephaleuros virescens is a pantropical subaerial green alga with no known long‐range dispersal mechanisms. Sexual reproduction is relatively rare and may involve intragametangial fusion of identical, mitotically produced gametes. This situation may be a consequence of adaptation to the subaerial habitat. Genetic variation among populations of C. virescens may be very low and might be positively correlated to the distance (hence, time) separating populations. Thus, assessing the global biogeography of C. virescens requires analysis of what might be low levels of variation. Because C. virescens occurs on literally hundreds of different host species, the question of host‐races must also be considered. Preliminary analysis of local populations of C. virescens, originally obtained as field collections from three different host species and subsequently raised in culture, is the first step in addressing the biogeography of this alga. We are using the AFLP plant mapping protocol by PE Applied Biosystems to detect genetic variability in the three isolates of C. virescens. AFLP is a PCR‐based DNA fingerprinting technique that detects the presence or absence of restriction fragments rather than fragment length differences. Because the number of restriction fragments that can be detected with the AFLP technique is “virtually unlimited,” it is a very powerful tool for assessing the degree of relatedness or variability among cultivars or isolates. AFLP techniques have been used successfully to distinguish morphologically identical bacteria, determine relatedness among soybean accessions, reveal genetic variability within bee samples, and identifyfall armyworm strains and hybrids.