Why clinical modulation of efflux transport at the human blood-brain barrier is unlikely: the ITC evidence-based position.

Drug interactions due to efflux transport inhibition at the blood-brain barrier (BBB) have been receiving increasing scrutiny because of the theoretical possibility of adverse central nervous system (CNS) effects identified in preclinical studies. In this review, evidence from pharmacokinetic, pharmacodynamic, imaging, pharmacogenetic, and pharmacovigilance studies, along with drug safety reports, is presented supporting a low probability of modulating transporters at the human BBB by currently marketed drugs.

The effect of changes in the health care environment on rehabilitation research: a survey of rehabilitation physicians.

OBJECTIVE: To assess what effect organizational, funding, and system changes in the health care environment may have on rehabilitation research. DESIGN: National survey. SETTING: Academic and clinical research programs. PARTICIPANTS: A total of 138 physicians participated in the survey. INTERVENTION: Mailed instrument requesting information on demographics, research activities, and indicators of change. MAIN OUTCOME MEASURES: Percentage of respondents reporting specific perceptions on (1) how academic and research programs are affected by organizational changes, (2) availability of research funds, and (3) the overall impact that health care changes have on research; between-group comparisons on survey responses. RESULTS: Usable responses were obtained from 138 physicians (response rate, 42.5%). Responding physicians reported workplace changes that included organizational restructuring (32.6%), affiliation with managed care plans (24%), and staff decreases (45.9%). Over half (54.8%) indicated that facility changes had detracted from their abilities to conduct research. A third (34.8%) reported declines in numbers of inpatient beds, and 89.6% reported decreased length of stay (LOS). Decreased LOS was cited as detracting from ongoing research by 36.6% and as discouraging new research by 33.3% of respondents. Although not reflected in measures of scholarly productivity, 53.6% reported having less time to devote to scholarship and 48.5% reported decreased professional activities. Over two thirds (67.4%) of responding physicians indicated that health care system changes had a negative impact on current research, and 54.5% indicated that such changes made it less likely that they would pursue new research. CONCLUSION: Changes in the health care system have had a dampening effect on rehabilitation physicians' research pursuits.

Fatty acid uptake and incorporation in brain: studies with the perfusion model.

The contributions of individual components of blood to brain [14C]palmitate uptake and incorporation were studied with the in situ brain perfusion technique in the pentobarbital-anesthetized rat. With whole-blood perfusate, brain unacylated [14C]palmitate uptake was linear with time and extrapolated to zero at T = 0 s of perfusion. Tracer accumulated in brain with a blood-to-brain transfer coefficient of 1.8 +/- 0.1 x 10(-4) mL/s/g (whole cerebral hemisphere). Incorporation into brain lipids was rapid such that approximately 40% of tracer in brain at 45 s of perfusion was in cerebral phospholipids and neutral lipids. Similar rates of uptake were obtained during unacylated [14C]palmitate perfusion in whole rat plasma, serum, or artificial saline containing 2-3% albumin, suggesting that albumin has a key role in determining [14C]palmitate uptake in brain. The excellent match in brain uptake rates between whole blood and albumin-containing saline fluid suggests that the perfusion technique will be useful method for quantifying the individual contributions of blood constituents and albumin binding on brain [14C]palmitate uptake.

Characterization of the blood-brain barrier choline transporter using the in situ rat brain perfusion technique.

Choline enters brain by saturable transport at the blood-brain barrier (BBB). In separate studies, both sodium-dependent and passive choline transport systems of differing affinity have been reported at brain capillary endothelial cells. In the present study, we re-examined brain choline uptake using the in situ rat brain perfusion technique. Saturable brain choline uptake from perfusion fluid was best described by a model with a single transporter (V:(max) = 2.4-3.1 nmol/min/g; K(m) = 39-42 microM) with an apparent affinity (1/Km)) for choline five to ten-fold greater than previously reported in vivo, but less than neuronal 'high-affinity' brain choline transport (K(m) = 1-5 microM). BBB choline uptake from a sodium-free perfusion fluid using sucrose for osmotic balance was 50% greater than in the presence of sodium suggesting that sodium is not required for transport. Hemicholinium-3 inhibited brain choline uptake with a K(i) (57 +/- 11 microM) greater than that at the neuronal choline system. In summary, BBB choline transport occurs with greater affinity than previously reported, but does not match the properties of the neuronal choline transporter. The V:(max) of this system is appreciable and may provide a mechanism for delivering cationic drugs to brain.

The effect of changes in the U.S. health care system on rehabilitation research: the results of a survey of rehabilitation health professionals.

To obtain empirical data on the impact of changes in the health care system on rehabilitation research, physicians, psychologists, nurses, physical therapists, and occupational therapists involved in such research were surveyed to determine: 1) how the changes had affected their research productivity; 2) whether their scholarly activities (e.g., publications) had been affected; and 3) whether working in an academic vs non-academic institution made a difference. There were 438 usable responses (38% response rate) to the survey, which was sent to members of national organizations. Although the results somewhat supported suggestions that changes in the system had had a dampening effect on research, they were ambiguous, failing to clearly demonstrate a negative impact of managed care. More rigorous study is needed to enable firm conclusions and the formulation of actions to address factors that may influence rehabilitation research.

Transport of glutamate and other amino acids at the blood-brain barrier.

In most regions of the brain, the uptake of glutamate and other anionic excitatory amino acids from the circulation is limited by the blood-brain barrier (BBB). In most animals, the BBB is formed by the brain vascular endothelium, which contains cells that are joined by multiple bands of tight junctions. These junctions effectively close off diffusion through intercellular pores; as a result, most solutes cross the BBB either by diffusing across the lipoid endothelial cell membranes or by being transported across by specific carriers. Glutamate transport at the BBB has been studied by both in vitro cell uptake assays and in vivo perfusion methods. The results demonstrate that at physiologic plasma concentrations, glutamate flux from plasma into brain is mediated by a high affinity transport system at the BBB. Efflux from brain back into plasma appears to be driven in large part by a sodium-dependent active transport system at the capillary abluminal membrane. Glutamate concentration in brain interstitial fluid is only a fraction of that of plasma and is maintained fairly independently of small fluctuations in plasma concentration. Restricted brain passage is also observed for several excitatory glutamate analogs, including domoic acid and kynurenic acid. In summary, the BBB is one component of a regulatory system that helps maintain brain interstitial fluid glutamate concentration independently of the circulation.

Tailored, interactive soap operas for breast cancer education of high-risk Hispanic women.

BACKGROUND: While Hispanic women have lower rates of breast cancer than do women of other ethnic groups, they are the least likely to undergo screening examinations. This study evaluated a culturally sensitive and linguistically appropriate, tailored, computer-based, educational program for early detection of breast cancer aimed at high-risk Hispanic women. METHODS: Spanish-speaking Hispanic women from an inner-city community health clinic were recruited and randomly assigned either to a computer intervention with an interactive soap-opera format (n = 118) or to a comparison group (n = 60). Pre- and posttests were used to identify any change in breast-cancer-related knowledge and beliefs. RESULTS: Both younger (18-40 years old) and older (41-65 years old) women in the intervention group demonstrated significant increases in their breast cancer screening knowledge and beliefs as compared with the younger and older women in the comparison group (n < 0.05). CONCLUSIONS: Computer-based tailored and interactive soap operas that are linguistically and culturally appropriate are effective in increasing breast cancer screening knowledge and beliefs among underserved Spanish-speaking Hispanic women.

Cerebrovascular Biology and the various neural barriers: challenges and future directions.

DESPITE MAJOR ADVANCES in neuroscience, potential therapeutic options for the treatment of central nervous system diseases often cannot be optimized secondary to the presence of the blood-brain barrier (BBB). During the next decade of inquiry, it is crucial that basic science and clinical research that is focused on overcoming the BBB, to optimize delivery to the central nervous system, be identified and supported as a priority topic. For this reason, the third international Cerebrovascular Biology and Blood-Brain Barrier Conference was convened in March 1998 in Gleneden Beach, OR. This meeting brought together basic science and clinical researchers from around the world to analyze BBB function and to discuss delivery of effective agents to the central nervous system for treatment of brain disease. This report summarizes the information presented at the meeting and the discussions that ensued. The current state of knowledge, obstacles to further understanding the BBB, and research priorities are identified.

Blood-brain barrier glucose transporter: effects of hypo- and hyperglycemia revisited.

The transport of glucose across the blood-brain barrier (BBB) is mediated by the high molecular mass (55-kDa) isoform of the GLUT1 glucose transporter protein. In this study we have utilized the tritiated, impermeant photolabel 2-N-[4-(1 -azi-2,2,2-trifluoroethyl)[2-3H]propyl]-1,3-bis(D-mannose-4-ylo xy)-2-propylamine to develop a technique to specifically measure the concentration of GLUT1 glucose transporters on the luminal surface of the endothelial cells of the BBB. We have combined this methodology with measurements of BBB glucose transport and immunoblot analysis of isolated brain microvessels for labeled luminal GLUT1 and total GLUT1 to reevaluate the effects of chronic hypoglycemia and diabetic hyperglycemia on transendothelial glucose transport in the rat. Hypoglycemia was induced with continuous-release insulin pellets (6 U/day) for a 12- to 14-day duration; diabetes was induced by streptozotocin (65 mg/kg i.p.) for a 14- to 21-day duration. Hypoglycemia resulted in 25-45% increases in regional BBB permeability-surface area (PA) values for D-[14C]glucose uptake, when measured at identical glucose concentration using the in situ brain perfusion technique. Similarly, there was a 23+/-4% increase in total GLUT1/mg of microvessel protein and a 52+/-13% increase in luminal GLUT1 in hypoglycemic animals, suggesting that both increased GLUT1 synthesis and a redistribution to favor luminal transporters account for the enhanced uptake. A corresponding (twofold) increase in cortical GLUT1 mRNA was observed by in situ hybridization. In contrast, no significant changes were observed in regional brain glucose uptake PA, total microvessel 55-kDa GLUT1, or luminal GLUT1 concentrations in hyperglycemic rats. There was, however, a 30-40% increase in total cortical GLUT1 mRNA expression, with a 96% increase in the microvessels. Neither condition altered the levels of GLUT3 mRNA or protein expression. These results show that hypoglycemia, but not hyperglycemia, alters glucose transport activity at the BBB and that these changes in transport activity result from both an overall increase in total BBB GLUT1 and an increased transporter concentration at the luminal surface.

Blood-brain barrier choline transport in the senescent rat.

Choline is an important membrane phospholipid constituent and a neurotransmitter precursor that is minimally synthesized in brain. The long-term maintenance of brain choline concentration is dependent on uptake from plasma, which occurs via saturable transporter at the blood-brain barrier. Previous studies have suggested that brain choline uptake declined with age. To reevaluate this, brain choline uptake in 3, 12, 24, and 28-month-old Fischer-344 rats was evaluated using the in situ brain perfusion technique. Minimal differences were found with uptake parameters differing by approximately 10% between aged and adult rats for tracer levels while similar trends were observed at higher choline concentrations. Further, estimated Vmax and Km values differed by <30% between the groups. The results suggest that blood-brain barrier choline uptake changes minimally with aging in the rat.

Training school counselors in substance abuse risk reduction techniques for use with children and adolescents.

A training project prepared school counselors for expanded roles in the prevention, early detection, and appropriate referral of students at high risk of substance abuse. The project trained middle and high school counselors to work as facilitators of support groups for students at greatest risk for substance abuse; the results were: 1) greater perceived self-efficacy, comfort, confidence, and competence by counselors as a result of Initial, Experiential, and Concurrent training, and 2) improved ability to use group counseling techniques as a result of training.

Beta-amyloid induced increase in choline flux across PC12 cell membranes.

Beta-amyloid peptide is the main constituent of senile plaques and is implicated in the pathogenesis of Alzheimer's disease. It has been shown to be both neurotoxic and neurotrophic in vivo, and its effects have been suggested to be mediated in part by alterations in membrane transport. In the present study, we investigated the effect of beta-amyloid (1-40) on choline transport in cultured PC12 cells. We found that exposure to 46 or 92 microM beta-amyloid (1-40) increased [14C]choline flux in PC12 cells in a concentration-dependent manner, whereas exposure to reverse sequence beta-amyloid (40-1) had no effect. If there is a similar effect in vivo, the increased beta-amyloid dependent permeability to choline could lead to depletion of cellular choline stores and could contribute to the selective vulnerability of cholinergic neurons in Alzheimer's disease.

In vivo imaging of fatty acid incorporation into brain to examine signal transduction and neuroplasticity involving phospholipids.

An in vivo method is presented that allows quantification and imaging of fatty acid incorporation into different brain phospholipids in relation to membrane synthesis, neuroplasticity, and signal transduction. The method can be used with positron emission tomography, and may help to evaluate brain phospholipid metabolism in humans with brain tumors, neurodegenerative disease, cerebral ischemia or trauma, or neurotoxic effects of drugs or other agents.

Subgingival temperature: relation to gingival crevicular fluid enzymes, cytokines, and subgingival plaque micro-organisms.

There have been no reports on the relationship of subgingival temperature to specific gingival crevicular fluid (GCF) components. Therefore, the purpose of this cross-sectional study was to determine whether there was any relationship between subgingival temperature and GCF levels of neutrophil elastase (NE), myeloperoxidase (MPO), beta-glucuronidase (BG), interleukin-1 alpha (IL-1), and interferon alpha (IFN). Furthermore, another objective was to confirm an association of subgingival temperature with clinical parameters and specific subgingival plaque micro-organisms as has been reported earlier. 27 human subjects each having healthy (n = 50), gingivitis (n = 59) and periodontitis (n = 53) sites were evaluated. The plaque index (PI), subgingival temperature, probing depth, attachment loss, bleeding index and gingival index were measured. GCF was sampled following the measurement of the PI and removal of the supragingival plaque. GCF samples were assayed for the enzymes NE, BG, MPO and the cytokines IFN-alpha and IL-1 alpha. A sterile Gracey curette was utilized at each sampled site to collect subgingival plaque. The plaque samples were evaluated using an immunoassay. Subgingival temperature was found to directly correlate with all clinical parameters (p < 0.001). Significant, albeit not large, correlations were found between subgingival temperature and NE (r = 0.35, p < 0.001), MPO (r = 0.26, p < 0.001) and BG (r = 0.23, p < 0.01). Temperature was found to correlate positively with E. corrodens (r = 0.33, p < 0.02) and F. nucleatum (r = 0.25, p < 0.05) but not with P. intermedia (r = 0.02, p = 0.9), P. gingivalis (r = 0.20, p = 0.1) and A. actinomycetemcomitans (r = 0.01, p > 0.9). In conclusion, subgingival temperature is correlated with the GCF enzymes, NE, MPO and BG as well as the clinical parameters and specific plaque micro-organisms associated with periodontal disease.

Facilitated brain uptake of 4-chlorokynurenine and conversion to 7-chlorokynurenic acid.

7-Chlorokynurenic acid (7-Cl-KYNA) and 5,7-dichlorokynurenic acid (5,7-Cl2-KYNA) are of therapeutic interest as potent glycine/N-methyl-D-aspartate NMDA) receptor antagonists, but are excluded from brain by the blood-brain barrier. We examined whether these compounds could be delivered to brain through their respective precursors, L-4-chlorokynurenine (4-Cl-KYN) and L-4,6-dichlorokynurenine (4,6-Cl2-KYN), which are amino acids. 4-Cl-KYN was shown to be rapidly shuttled into the brain by the large neutral amino acid transporter of the blood-brain barrier (K(m) = 105 +/- 14 microM, Vmax = 16.9 +/- 2.3 nmol min-1 g-1) and to be converted intracerebrally to 7-Cl-KYNA. 4,6-Cl2-KYN also expressed affinity for the transporter, but four-fold less than that of 4-Cl-KYN. In summary, the results show that because of their facilitated uptake 4-Cl-KYN and 4,6-Cl2KYN might be useful prodrugs for brain delivery of glycine-NMDA receptor antagonists.

Risk indicators for periodontal disease in a racially diverse urban population.

A cross-sectional study of 117 subjects from a dental clinic serving a diverse population (i.e., Whites, African-Americans, Native-Americans, and Asians) was performed to evaluate risk indicators of periodontal disease. Gingival crevicular fluid (GCF) and subgingival plaque were taken at the same visit from 4 posterior sites of the most diseased sextant in each subject. Age, smoking packyears, beta-glucuronidase (beta G), neutrophil elastase (NE), myeloperoxidase (MPO), Fusobacterium nucleatum (F. nucleatum), and Porphyromonas gingivalis (P. gingivalis) were significantly (p < 0.05-0.005) correlated with attachment loss. Probing depth was significantly correlated with smoking packyears, beta G, NE, MPO, F. nucleatum and Prevotella intermedia (P. intermedia) (p < 0.05-0.005). Mean NE value of Whites was lower than the mean NE values of African-Americans, Native-Americans and Asians (p < 0.05). Whites had a lower mean beta G value compared to African Americans, and a lower mean MPO value compared to African Americans and Native Americans. The %s of patients positive for F. nucleatum, P. intermedia and Eikenella corrodens (E. corrodens) were higher in Native Americans compared to Whites. Step-wise multiple regression analysis was performed to construct models for the estimation of probing depth and attachment loss. The most parsimonious regression models which had the best R2 values included the following variables and accounted for the indicated % of variability: models 1 and 2: beta G, race, and F. nucleatum accounted for 50% of the variability in mean probing depth and 39% of the variability in a single site (first molar) for probing depth, respectively; model 3: age, beta G, and F. nucleatum accounted for 53% of the variability in mean attachment loss; model 4: age, NE, and F. nucleatum explained 35% of the variability in a single site (first molar) for attachment loss. The results suggest that age, race, smoking packyears, beta G, NE, MPO, F. nucleatum, P. gingivalis and P. intermedia are risk indicators for periodontal disease in this racially diverse urban population. Regression models which include multiple variables (i.e., demographic factors, GCF enzymes and periodontopathic bacteria) can be used to estimate periodontal disease status.

Identification of two molecular species of rat brain phosphatidylcholine that rapidly incorporate and turn over arachidonic acid in vivo.

In vivo rates of arachidonic acid incorporation and turnover were determined for molecular species of rat brain phosphatidylcholine (PtdCho) and phosphatidylinositol (PtdIns). [3H]Arachidonic acid was infused intravenously in pentobarbital-anesthetized rats at a programmed rate to maintain constant plasma specific activity for 2-10 min. At the end of infusion, animals were killed by microwave irradiation, and brain phospholipids were isolated, converted to diacylglycerobenzoates, and resolved as molecular species by reversed-phase HPLC. Most [3H] arachidonate (> 87%) was incorporated into PtdCho and PtdIns, with arachidonic acid at the sn-2 position and with oleic acid (18:1), palmitic acid (16:0), or stearic acid (18:0) at the sn-1 position. However, 10-15% of labeled brain PtdCho eluted in a small peak containing two molecular species with arachidonic acid at the sn-2 position and palmitoleic acid (16:1) or linoleic acid (18:2) at the sn-1 position. Analysis demonstrated that tracer was present in both the 16:1-20:4 and 18:2-20:4 PtdCho species at specific activities 10-40 times that of the other phospholipids. Based on the measured mass of arachidonate in each phospholipid molecular species, half-lives were calculated for arachidonate of < 10 min in 16:1-20:4 and 18:2-20:4 PtdCho and 1-3 h in 16:0-20:4, 18:1-20:4 PtdCho and PtdIns. The very short half-lives for arachidonate in the 16:1-20:4 and 18:2-20:4 PtdCho molecular species suggest important roles for these molecules in brain phospholipid metabolism and signal transduction.

Crevicular fluid enzymes from endosseous dental implants and natural teeth.

Several neutrophil-derived enzymes that are present in the gingival crevicular fluid have been evaluated for use as risk markers for periodontal disease progression. However, very little information is available about the presence of these enzymes in peri-implant tissues. The purpose of this cross-sectional study was to compare levels of enzymes in gingival crevicular fluid between natural teeth and endosseous dental implants and between well-integrated and failing implants. Scores of plaque and gingivitis were recorded for 68 integrated implants, five failing implants, and 34 natural teeth in 12 completely edentulous and 18 partially edentulous subjects. Samples of gingival crevicular fluid were obtained from these sites using filter paper strips and were assayed for levels of neutral protease, neutrophil elastase, myeloperoxidase, and beta-glucuronidase. Neutral protease levels were higher (P = .066) at moderately to severely inflamed implant sites (Gingival Index of 2, 3) compared to mildly or noninflamed sites (Gingival Index of = 0, 1). Despite the small number (n = 5) of failing implants evaluated in this study, levels of neutrophil elastase, myeloperoxidase, and beta-glucuronidase were significantly higher (P < or = .001) around failing implants compared to successful implants. Neutral protease levels were also elevated around failing implants, but the difference was not statistically significant. Results of this study indicate that neutrophil elastase, myeloperoxidase, and beta-glucuronidase levels in GCF appear to be good candidates for study as risk markers of implant failure.