Showing papers by "University of California, San Francisco published in 1989"

Reversible middle cerebral artery occlusion without craniectomy in rats.

Abstract: To develop a simple, relatively noninvasive small-animal model of reversible regional cerebral ischemia, we tested various methods of inducing infarction in the territory of the right middle cerebral artery (MCA) by extracranial vascular occlusion in rats. In preliminary studies, 60 rats were anesthetized with ketamine and different combinations of vessels were occluded; blood pressure and arterial blood gases were monitored. Neurologic deficit, mortality rate, gross pathology, and in some instances, electroencephalogram and histochemical staining results were evaluated in all surviving rats. The principal procedure consisted of introducing a 4-0 nylon intraluminal suture into the cervical internal carotid artery (ICA) and advancing it intracranially to block blood flow into the MCA; collateral blood flow was reduced by interrupting all branches of the external carotid artery (ECA) and all extracranial branches of the ICA. In some groups of rats, bilateral vertebral or contralateral carotid artery occlusion was also performed. India ink perfusion studies in 20 rats documented blockage of MCA blood flow in 14 rats subjected to permanent occlusion and the restoration of blood flow to the MCA territory in six rats after withdrawal of the suture from the ICA. The best method of MCA occlusion was then selected for further confirmatory studies, including histologic examination, in five additional groups of rats anesthetized with halothane. Seven of eight rats that underwent permanent occlusion of the MCA had resolving moderately severe neurologic deficits (Grade 2 of 4) and unilateral infarcts averaging 37.6 +/- 5.5% of the coronal sectional area at 72 hours after the onset of occlusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Functional status and well-being of patients with chronic conditions. Results from the Medical Outcomes Study.

Abstract: Enhancing daily functioning and well-being is an increasingly advocated goal in the treatment of patients with chronic conditions. We evaluated the functioning and well-being of 9385 adults at the time of office visits to 362 physicians in three US cities, using brief surveys completed by both patients and physicians. For eight of nine common chronic medical conditions, patients with the condition showed markedly worse physical, role, and social functioning; mental health; health perceptions; and/or bodily pain compared with patients with no chronic conditions. Each condition had a unique profile among the various health components. Hypertension had the least overall impact; heart disease and patient-reported gastrointestinal disorders had the greatest impact. Patients with multiple conditions showed greater decrements in functioning and well-being than those with only one condition. Substantial variations in functioning and well-being within each chronic condition group remain to be explained. (JAMA. 1989;262:907-913)

Induction of angiogenesis during the transition from hyperplasia to neoplasia

Abstract: It is now well established that unrestricted growth of tumours is dependent upon angiogenesis. Previous studies on tumour growth, however, have not revealed when or how the transition to an angiogenic state occurs during early tumour development. The advent of transgenic mice carrying oncogenes that reproducibly elicit tumours of specific cell types is providing a new format for studying multi-step tumorigenesis. In one of these models, transgenic mice expressing an oncogene in the beta-cells of the pancreatic islets heritably recapitulate a progression from normality to hyperplasia to neoplasia. We report here that angiogenic activity first appears in a subset of hyperplastic islets before the onset of tumour formation. A novel in vitro assay confirms that hyperplasia per se does not obligate angiogenesis. Rather, a few hyperplastic islets become angiogenic in vitro at a time when such islets are neovascularized in vivo and at a frequency that correlates closely with subsequent tumour incidence. These findings suggest that induction of angiogenesis is an important step in carcinogenesis.

Risk factors for recurrent nonsyncopal falls. A prospective study.

Abstract: Falls are a major threat to the health of older persons. We evaluated potential risk factors for falls in 325 community-dwelling persons aged 60 years or older who had fallen during the previous year, then followed up weekly for 1 year to ascertain nonsyncopal falls and their consequences. Risk factors for having a single fall were few and relatively weak, but multiple falls were more predictable. In multivariate analyses, we found increased odds of two or more falls for persons who had difficulty standing up from a chair, difficulty performing a tandem walk, arthritis, Parkinson's disease, three or more falls during the previous year, and a fall with injury during the previous year, and for whites. The proportion of subjects with two or more falls per year increased from 0.10 for those with none or one of these risk factors to 0.69 for those with four or more risk factors. Among older persons with a history of a recent fall, the risk of multiple nonsyncopal falls can be predicted from a few simple questions and examinations. ( JAMA . 1989;261:2663-2668)

Targeted sampling: options for the study of hidden populations

Abstract: This paper describes some of the efforts of an interdisciplinary research team investigating the transmission of human immunodeficiency virus (HIV), the causative pathogen associated with the acquired immunodeficiency syndrome (AIDS) and related conditions. The risk groups studied were injecting drug users and their sexual partners. Due to the clandestine nature of illicit drug use, we were faced with two interrelated problems: developing a scientific method to monitor the spread of the HIV infection among these drug users and their sexual partners, groups generally thought to be especially difficult to reach; and creating a health education intervention that would help stop the epidemic from spreading among this population and through them to other members of the community. The method we developed to sample injecting drug users is called targeted sampling. Although it incorporates some aspects of other well established sampling strategies, it is sufficiently different to be treated as a separate research method. Further, targeted sampling provides a cohesive set of research methods that can help researchers study health or social problems that exist among populations that are difficult to reach because of their attributed social stigma, legal status, and consequent lack of visibility.

The role of excitatory amino acids and NMDA receptors in traumatic brain injury.

Abstract: Brain injury induced by fluid percussion in rats caused a marked elevation in extracellular glutamate and aspartate adjacent to the trauma site. This increase in excitatory amino acids was related to the severity of the injury and was associated with a reduction in cellular bioenergetic state and intracellular free magnesium. Treatment with the noncompetitive N-methyl-D-aspartate (NMDA) antagonist dextrophan or the competitive antagonist 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid limited the resultant neurological dysfunction; dextrorphan treatment also improved the bioenergetic state after trauma and increased the intracellular free magnesium. Thus, excitatory amino acids contribute to delayed tissue damage after brain trauma; NMDA antagonists may be of benefit in treating acute head injury.

Reciprocal expression of interferon gamma or interleukin 4 during the resolution or progression of murine leishmaniasis. Evidence for expansion of distinct helper T cell subsets.

Abstract: We purified poly(A)+ mRNA from the spleen and lymph nodes at designated times after infection with Leishmania major in genetically susceptible BALB/c and resistant C57BL/6 mice. The steady-state levels of IL-2, IFN-gamma, IL-4, and IL-1 beta mRNA were determined using Northern hybridizations. IL-2 mRNA levels in the infected organs of BALB/c and C57BL/6 mice were comparable after infection, but IFN-gamma and IL-4 mRNA levels were reciprocally expressed. Levels of IFN-gamma mRNA in C57BL/6 draining nodes and spleen were significantly greater than in BALB/c mice except at 4 and 6 wk of infection, when splenic IFN-gamma mRNA levels were transiently comparable. In contrast, IL-4 mRNA was apparent only in BALB/c and not in C57BL/6 nodes and spleen. Tissue levels of IL-1 beta mRNA were 10-20-fold greater in BALB/c mice. BALB/c mice were pretreated with GK1.5 mAb, a manipulation that promotes healing of subsequent infection by transiently depleting L3T4+ cells. At 8 wk of infection, by which time lymphoid organs were repopulated with L3T4+ cells, GK1.5-pretreated BALB/c mice produced IFN-gamma, but not IL-4 message. Serum levels of IgE were markedly elevated in infected BALB/c, but not in infected C57BL/6 or GK1.5-pretreated BALB/c mice, consistent with in vivo biologic activity of IL-4 in nonhealing mice. Treatment of infected BALB/c mice with neutralizing anti-IL-4 antibody abolished the elevation of serum IgE and significantly attenuated the progression of disease as assessed by size and ulceration of the lesion, and by reduction in the number of tissue parasites. Both protective and deleterious responses to Leishmania infection have previously been shown to be L3T4+ cell dependent. Our findings are consistent with the differential expansion of protective, IFN-gamma-producing Th1 cells in healing mice, and the expansion of deleterious, IL-4-producing Th2 cells in nonhealing mice. The inverse relationship of IFN-gamma and IL-4 gene expression during leishmaniasis may underlie the divergence of cellular and humoral immunity that occurs during chronic infection with Leishmania and possibly other intracellular parasites.

A controlled trial of leuprolide with and without flutamide in prostatic carcinoma.

Abstract: To test the hypothesis that maximal androgen blockade improves the effectiveness of the treatment of prostatic cancer, we conducted a randomized, double-blind trial in patients with disseminated, previously untreated prostate cancer (stage D2). All 603 men received leuprolide, an analogue of gonadotropin-releasing hormone that inhibits the release of gonadotropins, in combination with either placebo or flutamide, a nonsteroidal antiandrogen that inhibits the binding of androgens to the cell nucleus. As compared with the 300 patients receiving leuprolide and placebo, the 303 patients randomly assigned to receive leuprolide and flutamide had a longer progression-free survival (16.5 vs. 13.9 months; P = 0.039) and an increase in the median length of survival (35.6 vs. 28.3 months; P = 0.035). The differences between the treatments were particularly evident for men with minimal disease and good performance status; however, further studies should be conducted in this subgroup. Symptomatic improvement was greatest during the first 12 weeks of the combined androgen blockade, when leuprolide alone often produces a painful flare in the disease. We conclude that in patients with advanced prostate cancer, treatment with leuprolide and flutamide is superior to treatment with leuprolide alone.

Attachment of a 40-base-pair G + C-rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain reaction results in improved detection of single-base changes.

Abstract: Denaturing gradient gel electrophoresis (DGGE) can be used to distinguish two DNA molecules that differ by as little as a single-base substitution. This method detects approximately 50% of all possible single-base changes in DNA fragments ranging from 50 to approximately 1000 base pairs. To increase the number of single-base changes that can be distinguished by DGGE, we used the polymerase chain reaction to attach a 40-base-pair G + C-rich sequence, designated a GC-clamp, to one end of amplified DNA fragments that encompass regions of the mouse and human beta-globin genes. We show that this GC-clamp allows the detection of mutations, including the hemoglobin sickle (HbS) and hemoglobin C (HbC) mutations within the human beta-globin gene, that were previously indistinguishable by DGGE. In addition to providing an easy way to attach a GC-clamp to genomic DNA fragments, the polymerase chain reaction technique greatly increases the sensitivity of DGGE. With this approach, DNA fragments derived from less than 5 ng of human genomic DNA can be detected by ethidium bromide staining of the gel, obviating the need for radioactive probes. These improvements extend the applicability of DGGE for the detection of polymorphisms and mutations in genomic and cloned DNA.

GTPase inhibiting mutations activate the α chain of G s and stimulate adenylyl cyclase in human pituitary tumours

Abstract: A subset of growth hormone-secreting human pituitary tumours carries somatic mutations that inhibit GTPase activity of a G protein alpha chain, alpha(s) The resulting activation of adenylyl cyclase bypasses the cells' normal requirement for trophic hormone Amino acids substituted in the putative gsp oncogene identify a domain of G protein alpha-chains required for intrinsic ability to hydrolyse GTP This domain may serve as a built-in counter-part of the separate GTPase-activating proteins required for GTP hydrolysis by small GTP-binding proteins such as p21ras

Cyclin synthesis drives the early embryonic cell cycle.

Abstract: We have produced extracts of frog eggs that can perform multiple cell cycles in vitro. Destruction of the endogenous messenger RNA arrests the extracts in interphase. The addition of exogenous cyclin mRNA is sufficient to produce multiple cell cycles. The newly synthesized cyclin protein accumulates during each interphase and is degraded at the end of each mitosis.

Signal transduction through the fibronectin receptor induces collagenase and stromelysin gene expression.

Abstract: We have investigated the effects of ligation of the fibronectin receptor (FnR) on gene expression in rabbit synovial fibroblasts. Monoclonal antibodies to the FnR that block initial adhesion of fibroblasts to fibronectin induced the expression of genes encoding the secreted extracellular matrix-degrading metalloproteinases collagenase and stromelysin. That induction was a direct consequence of interaction with the FnR was shown by the accumulation of mRNA for stromelysin and collagenase. Monoclonal antibodies to several other membrane glycoprotein receptors had no effect on metalloproteinase gene expression. Less than 2 h of treatment of the fibroblasts with anti-FnR in solution was sufficient to trigger the change in gene expression, and induction was blocked by dexamethasone. Unlike other inducers of metalloproteinase expression, including phorbol diesters and growth factors, addition of the anti-FnR in solution to cells adherent to serum-derived adhesion proteins or collagen produced no detectable change in cell shape or actin microfilament organization. Inductive effects were potentiated by cross-linking of the ligand. Fab fragments of anti-FnR were ineffective unless cross-linked or immobilized on the substrate. Adhesion of fibroblasts to native fibronectin did not induce metallo-proteinases. However, adhesion to covalently immobilized peptides containing the arg-gly-asp sequence that were derived from fibronectin, varying in size from hexapeptides up to 120 kD, induced collagenase and stromelysin gene expression. This suggests that degradation products of fibronectin are the natural inductive ligands for the FnR. These data demonstrate that signals leading to changes in gene expression are transduced by the FnR, a member of the integrin family of extracellular matrix receptors. The signaling of changes in gene expression by the FnR is distinct from signaling involving cell shape and actin cytoarchitecture. At least two distinct signals are generated: the binding of fibronectin-derived fragments and adhesion-blocking antibodies to the FnR triggers events different from those triggered by binding of the native fibronectin ligand. Because the genes regulated by this integrin are for enzymes that degrade the extracellular matrix, these results suggest that information transduced by the binding of various ligands to integrins may orchestrate the expression of genes regulating cell behavior in the extracellular environment.

The role of cyclin synthesis and degradation in the control of maturation promoting factor activity

Abstract: We show that cyclin plays a pivotal role in the control of mitosis. A proteolysis-resistant mutant of cyclin prevents the inactivation of maturation promoting factor and the exit from mitosis both in vivo and in vitro. We have used a fractionated extract to study the activation of MPF by added cyclin protein.

An essential role for postsynaptic calmodulin and protein kinase activity in long-term potentiation

Abstract: THE phenomenon of long-term potentiation (LTP), a long lasting increase in the strength of synaptic transmission which is due to brief, repetitive activation of excitatory afferent fibres, is one of the most striking examples of synaptic plasticity in the mammalian brain. In the CA1 region of the hippocampus, the induction of LTP requires activation of NMDA (N-methyl-D-aspartate) receptors by synaptically released glutamate1 with concomitant postsynaptic membrane depolarization2-5. This relieves the voltage-dependent magnesium block of the NMD A-receptor ion channel6,7, allowing calcium to flow into the dendritic spine8-10. Although calcium has been shown to be a necessary trigger for LTP (refs 11,12), little is known about the immediate biochemical processes that are activated by calcium and are responsible for LTP. The most attractive candidates have been calcium/cal-modulin-dependent protein kinase II (CaM-KII) (refs 13-16), protein kinase C (refs 17-19), and the calcium-dependent protease, calpain20. Extracellular application of protein kinase inhibitors to the hippocampal slice preparation blocks the induction of LTP (refs 21-23) but it is unclear whether this is due to a pre- and/or postsynaptic action. We have found that intracellular injection into CA1 pyramidal cells of the protein kinase inhibitor H-7, or of the calmodulin antagonist calmidazolium, blocks LTP. Further-more, LTP is blocked by the injection of synthetic peptides that are potent calmodulin antagonists and inhibit CaM-KII auto- and substrate phosphorylation. These findings demonstrate that in the postsynaptic cell both activation of calmodulin and kinase activity are required for the generation of LTP, and focus further attention on the potential role of CaM-KII in LTP.

Movement of microtubules by single kinesin molecules

Abstract: Kinesin is a motor protein that uses energy derived from ATP hydrolysis to move organelles along microtubules. Using a new technique for measuring the movement produced in vitro by individual kinesin molecules, it is shown that a single kinesin molecule can move a microtubule for several micrometers. New information about the mechanism of force generation by kinesin is presented.

Linkage of a prion protein missense variant to Gerstmann–Sträussler syndrome

Abstract: Gerstmann-Straussler syndrome is a rare familial neurodegenerative condition that is vertically transmitted, in an apparently autosomal dominant way. It can also be horizontally transmitted to non-human primates and rodents through intracerebral inoculation of brain homogenates from patients with the disease. The exact incidence of the syndrome is unknown but is estimated to be between one and ten per hundred million. Patients initially suffer from ataxia or dementia and deteriorate until they die, in one to ten years. Protease-resistant prion protein (PrP) and PrP-immunoreactive amyloid plaques with characteristic morphology accumulate in the brains of these patients. Current diagnostic criteria for Gerstmann-Straussler syndrome incorporate clinical and neuropathological features, as animal transmission studies can be unreliable. PrP is implicated in the pathogenesis and transmission of the condition and in scrapie, an equivalent animal disease. It was discovered by enriching scrapie-infected hamster brain fractions for infectivity. Because there is compelling evidence that the scrapie isoform of PrP is a necessary component of the infectious particle, it seemed possible that the PrP gene on the short arm of human chromosome 20 in Gerstmann-Straussler syndrome might be abnormal. We show here that PrP codon 102 is linked to the putative gene for the syndrome in two pedigrees, providing the best evidence to date that this familial condition is inherited despite also being infectious, and that substitution of leucine for proline at PrP codon 102 may lead to the development of Gerstmann-Straussler syndrome.

Dominoes and clocks: the union of two views of the cell cycle.

Abstract: We review the recent advances in understanding transitions within the cell cycle. These have come from both genetic and biochemical approaches. We discuss the phylogenetic conservation of the mechanisms that induce mitosis and their implications for other transitions in the cell cycle.

Fluorescence microscopy in three dimensions.

Abstract: The combination of the specificity provided by fluorescence microscopy and the ability to quantitatively analyze specimens in three dimensions allows the fundamental organization of cells to be probed as never before Key features in this emergent technology have been the development of a wide variety of fluorescent dyes or fluorescently labeled probes to provide the requisite specificity High-quality, cooled charge-coupled devices have recently become available Functioning as nearly ideal imagers or "electronic film," they are more sensitive than photomultipliers and provide extraordinarily accurate direct digital readout from the microscope Not only is this precision crucial for accurate quantitative imaging such as that required for the ratioing necessary to determine intracellular ion concentrations, but it also opens the way for sophisticated image processing It is important to realize that image processing isn't simply a means to improve image aesthetics, but can directly provide new, biologically important information The impact of modern video microscopy techniques (Allen, 1985; Inoue, 1986) attests to the fact that many biologically relevant phenomena take place at the limits of conventional microscopy Image processing can be used to substantially enhance the resolution and contrast obtainable in two dimensions, enabling the invisible to be seen and quantitated Cells are intrinsically three-dimensional This can simply be a nuisance because of limited depth of focus of the microscope or it could be a fundamental aspect of the problem being studied In either case, image processing techniques can be used to rapidly provide the desired representation of the data In this chapter we have discussed the nature of image formation in three dimensions and dealt with several means to remove contaminating out-of-focus information The most straightforward of these methods uses only information from adjacent focal planes to correct the central one This approach can be readily applied to virtually any problem and with most commonly available image processing hardware to provide a substantially deblurred image in almost real time In addition to covering more sophisticated algorithms where the utmost in three-dimensional imaging is required, we have developed a method for extremely rapidly and accurately producing an in-focus, high-resolution "synthetic projection" image from a thick specimen This is equivalent to that produced by a microscope having the impossible combination of a high-NA objective lens and an infinite depth of focus A variation on this method allows efficient calculation of stereo pairs(ABSTRACT TRUNCATED AT 400 WORDS)

Lifetime Risks of Hip, Colles', or Vertebral Fracture and Coronary Heart Disease Among White Postmenopausal Women

Abstract: • Lifetime risk is a useful way to estimate and compare the risk of various conditions. Hip fractures, Colles' fractures, and coronary heart disease, and breast and endometrial cancers are important conditions in postmenopausal women that might be influenced by the use of hormone replacement therapy. We used population-based data to estimate a woman's lifetime risk of suffering a hip, Colles', or vertebral fracture and her risk of dying of coronary heart disease. A 50-year-old white woman has a 16% risk of suffering a hip fracture, a 15% risk of suffering a Colles' fracture, and a 32% risk of suffering a vertebral fracture during her remaining lifetime. These risks exceed her risk of developing breast or endometrial cancer. She has a 31% risk of dying of coronary heart disease, which is about 10 times greater than her risk of dying of hip fractures or breast cancer. These lifetime risks provide a useful description of the comparative risks of conditions that might be influenced by postmenopausal hormone therapy. ( Arch Intern Med. 1989;149:2445-2448)

Ocular dominance column development: analysis and simulation

Abstract: The visual cortex of many adult mammals has patches of cells that receive inputs driven by the right eye alternating with patches that receive inputs driven by the left eye. These ocular dominance patches (or "columns") form during early life as a consequence of competition between the activity patterns of the two eyes. A mathematical model of several biological mechanisms that can account for this development is presented. Analysis of this model reveals the conditions under which ocular dominance segregation will occur and determines the resulting patch width. Simulations of the model also exhibit other phenomena associated with early visual development, such as topographic refinement of cortical receptive fields, the confinement of input cell connections to patches, monocular deprivation plasticity including a critical period, and the effect of artificially induced strabismus. The model can be used to predict the results of proposed experiments and to discriminate among various mechanisms of plasticity.

Treatment of Wilms' tumor. Results of the Third National Wilms' Tumor Study.

Abstract: The Third National Wilms' Tumor Study sought to reduce treatment for low-risk patients and find better chemotherapy for those at high risk for relapse. Eligible patients (1439) were randomized according to stage (I-IV) and histology (favorable [FH] or unfavorable [UH]), and contributed data to survival and relapse-free survival (RFS) analyses. Four-year (postnephrectomy) survival percentages and randomized treatment regimens for low-risk patients were 96.5% for 607 Stage I/FH patients who received dactinomycin (Actinomycin D [AMD], Merck Sharp & Dohme, West Point, PA) and vincristine (VCR) for 10 weeks versus 6 months; 92.2% for 278 Stage II/FH patients; and 86.9% for 275 Stage III/FH patients who received AMD + VCR +/- Adriamycin (ADR, Adria Laboratories, Columbus, OH) for 15 months. Stage II/FH patients also had either zero or 2000 cGy irradiation (RT) postoperatively and Stage III/FH patients either 1000 or 2000 cGy. Four-year survival was 73.0% for 279 high-risk patients (any Stage IV, all UH) who received postoperative radiation therapy (RT) and AMD + VCR + ADR +/- cyclophosphamide (CPM). Statistical analysis of survival and RFS experience shows that the less intensive therapy does not worsen results for low-risk patients and CPM does not benefit those at high risk.

Purification and complementary DNA cloning of a receptor for basic fibroblast growth factor

Abstract: Basic fibroblast growth factor (bFGF) participates in many processes including early developmental events, angiogenesis, wound healing, and maintenance of neuronal cell viability. A 130-kilodalton protein was isolated on the basis of its ability to specifically bind to bFGF. A complementary DNA clone was isolated with an oligonucleotide probe corresponding to determined amino acid sequences of tryptic peptide fragments of the purified protein. The putative bFGF receptor encoded by this complementary DNA is a transmembrane protein that contains three extracellular immunoglobulin-like domains, an unusual acidic region, and an intracellular tyrosine kinase domain. These domains are arranged in a pattern that is different from that of any growth factor receptor described.

Association between a specific apolipoprotein B mutation and familial defective apolipoprotein B-100.

Abstract: Familial defective apolipoprotein (apo) B-100 is a genetic disease that leads to hypercholesterolemia and to an increased serum concentration of low density lipoproteins that bind defectively to the apoB,E(LDL) receptor. The disorder appears to result from a mutation in the gene for apoB-100. Extensive sequence analysis of the two alleles of one subject heterozygous for the disorder has revealed a previously unreported mutation in the codon for amino acid 3500 that results in the substitution of glutamine for arginine. This same mutant allele occurs in six other, unrelated subjects and in eight affected relatives in two of these families. A partial haplotype of this mutant apoB-100 allele was constructed by sequence analysis and restriction enzyme digestion at positions where variations in the apoB-100 are known to occur. This haplotype is the same in three probands and four affected members of one family and lacks a polymorphic Xba I site whose presence has been correlated with high cholesterol levels. Thus, it appears that the mutation in the codon for amino acid 3500 (CGG----CAG), a CG mutational "hot spot," defines a minor apoB-100 allele associated with defective low density lipoproteins and hypercholesterolemia.

Polewards microtubule flux in the mitotic spindle: evidence from photoactivation of fluorescence.

Abstract: I have synthesized a novel derivative of carboxyfluorescein that is nonfluorescent, but can be converted to a fluorescent form by exposure to 365-nm light. This photoactivable, fluorescent probe was covalently attached to tubulin and microinjected into mitotic tissue culture cells, where it incorporated into functional spindles. To generate a fluorescent bar across the mitotic spindle, metaphase cells were irradiated with a slit microbeam. This bar decreased in intensity over the first minute, presumably due to turnover of nonkinetochore microtubules. The remaining fluorescent zones, now presumably restricted to kinetochore microtubules, moved polewards at 0.3-0.7 microns/min. This result provides strong evidence for polewards flux in kinetochore microtubules. In conjunction with earlier biotin-tubulin incorporation experiments (Mitchison, T. J., L. Evans, E. Schulze, and M. Kirschner. 1986. Cell. 45:515-527), I conclude that microtubules polymerize at kinetochores and depolymerize near the poles throughout metaphase. The significance of this observation for spindle structure and function is discussed. Local photoactivation of fluorescence should be a generally useful method for following molecular dynamics inside living cells.