Learning histology - dental and medical students' study strategies.

PURPOSE: Histology, the science of cells and tissues at the microscopic level, is an integral component of most dental and medical curricula and is often taught using both traditional and novel computer-based didactic approaches. The purpose of this study was to analyse the strategies used by dental and medical students when studying this very visual and challenging subject. METHODS: Data were collected from 75 dental and 143 medical students, who had almost identical histology learning resources at their disposal. RESULTS: When compared with their medical counterparts, dental students view histology as a more difficult subject and as less relevant for their future career. Whereas dental students, who are required to attend class unlike medical students, made more use of in-classroom learning opportunities, they did not take as much advantage of out-of-classroom resources. In addition, dental students reported a significantly higher tendency than medical students to work together, rather than to study alone. DISCUSSION: Small differences in the dental versus the medical learning environment associate with several observed differences in learning strategies that are adopted by dental and medical students. CONCLUSIONS: These differences should be considered when teaching the subject of histology to dental or to medical students.

Clostridium botulinum neurotoxins act with a wide range of potencies on SH-SY5Y human neuroblastoma cells.

We have described, in undifferentiated SH-SY5Y human neuroblastoma cells, the relative potency of Clostridium botulinum neurotoxin (BoNT) serotypes A-F Sensitivity of stimulated [3H]-noradrenaline ([3H]-NA) release to the toxins had a rank order of potency of: C > D > A > B > F after 3 days exposure. The difference between the most potent (BoNT/C: IC50 0.54 nM) and the least (BoNT/F: IC50 > 300 nM) was approximately 1,000-fold. Though fluid phase endocytosis may have been the mechanism of entry for low potency toxins the far higher potency of BoNT/C would suggest receptor-driven entry. Potency was not a reflection of the dependence of the release mechanism on a particular SNARE since the substrate specificities were mixed throughout the potency order. This indicated that the toxins differed in their efficiency of binding/endocytosis or enzymatic activity inside the cell. The serotypes that cleaved vesicle-associated membrane protein (VAMP) isoforms (BoNT/B, D and F) did not fully inhibit [3H]-NA release. Cleavage of the appropriate substrate proteins was observed for all serotypes. SNAP-25 cleavage by BoNT/A was shown to be a dose-dependent and correlated closely with reduction of release, supporting proteolysis as the mechanism by which toxin inhibited secretion. Comparison of the SH-SY5Y cell line sensitivity to BoNT/A with glycine releasing rat primary spinal cord neuron cultures, revealed a massive difference in potency; the primary cultures being approximately 200,000-fold more sensitive. The demonstration, using BoNTs, of the crucial role of SNAP-25, VAMP and syntaxin in SH-SY5Y cells suggests the use of this neuroblastoma as a model in the study of these proteins in neurotransmitter release.

A conjugate composed of nerve growth factor coupled to a non-toxic derivative of Clostridium botulinum neurotoxin type A can inhibit neurotransmitter release in vitro.

Nerve growth factor (NGF) receptor binding, internalisation and transportation of NGF has been identified as a potential route of delivery for other molecules. A derivative of Clostridium botulinum neurotoxin type A (LHN) that retains catalytic activity but has significantly reduced cell-binding capability has been prepared and chemically coupled to NGF. Intact clostridial neurotoxins potently inhibit neurotransmitter release at the neuromuscular junction by proteolysis of specific components of the vesicle docking/fusion complex. Here we report that the NGF-LHN/A conjugate, when applied to PC12 cells, significantly inhibited neurotransmitter release and cleaved the type A toxin substrate. This work represents the successful use of NGF as a targeting moiety for the delivery of a neurotoxin fragment.

Inhibition of vesicular secretion in both neuronal and nonneuronal cells by a retargeted endopeptidase derivative of Clostridium botulinum neurotoxin type A.

Clostridial neurotoxins potently and specifically inhibit neurotransmitter release in defined cell types by a mechanism that involves cleavage of specific components of the vesicle docking/fusion complex, the SNARE complex. A derivative of the type A neurotoxin from Clostridium botulinum (termed LH(N)/A) that retains catalytic activity can be prepared by proteolysis. The LH(N)/A, however, lacks the putative native binding domain (H(C)) of the neurotoxin and is thus unable to bind to neurons and effect inhibition of neurotransmitter release. Here we report the chemical conjugation of LH(N)/A to an alternative cell-binding ligand, wheat germ agglutinin (WGA). When applied to a variety of cell lines, including those that are ordinarily resistant to the effects of neurotoxin, WGA-LH(N)/A conjugate potently inhibits secretory responses in those cells. Inhibition of release is demonstrated to be ligand mediated and dose dependent and to occur via a mechanism involving endopeptidase-dependent cleavage of the natural botulinum neurotoxin type A substrate. These data confirm that the function of the H(C) domain of C. botulinum neurotoxin type A is limited to binding to cell surface moieties. The data also demonstrate that the endopeptidase and translocation functions of the neurotoxin are effective in a range of cell types, including those of nonneuronal origin. These observations lead to the conclusion that a clostridial endopeptidase conjugate that can be used to investigate SNARE-mediated processes in a variety of cells has been successfully generated.

Sensitivity of embryonic rat dorsal root ganglia neurons to Clostridium botulinum neurotoxins.

Clostridium botulinum neurotoxins (BoNT) are zinc dependent endopeptidases which, once internalised into the neuronal cytosol, block neurotransmission by proteolysis of membrane-associated proteins putatively involved in synaptic vesicle docking and fusion with the plasma membrane. Although many studies have used a variety of cellular systems to study the neurotoxins, most require relatively large amounts of toxin or permeabilisation to internalise the neurotoxin. We present here a primary culture of embryonic rat dorsal root ganglia (DRG) neurons that exhibits calcium-dependent substance P secretion when depolarised with elevated extracellular potassium and is naturally BoNT sensitive. The DRG neurons showed a different IC50 for each of the toxins tested with a 1000 fold difference between the most and least potent neurotoxins (0.05, 0.3, 30 and approximately 60 nM for A, C, F and B, respectively). BoNT/A cleavage of SNAP-25 was seen as early as 2 h, but substance P secretion was not significantly inhibited until 4 h intoxication and the effects of BoNT/A were observed for as long as 15 days. This primary neuronal culture system represents a new and sensitive cellular model for the in vitro study of the botulinum neurotoxins.

Capsaicin-stimulated release of substance P from cultured dorsal root ganglion neurons: involvement of two distinct mechanisms.

Capsaicin, the pungent component of "hot" chili peppers, selectively activates a distinct population of primary sensory neurons responsive to noxious stimuli. Many of these fibres express neuropeptides including the tachykinin, substance P. Using cultured dorsal root ganglion neurons, we found that capsaicin (10 microM) stimulated a 2-fold increase in release of substance P in the absence of extracellular Ca(2+). Elevated potassium (75 mM) was unable to induce release under these conditions. The introduction of Ca(2+) enhanced capsaicin-induced release and brought about a robust response to potassium. Preincubation of cells with botulinum neurotoxin A (100 nM) completely blocked potassium-induced release but the capsaicin response, in the absence of Ca(2+), was unaffected. However, toxin treatment dramatically reduced capsaicin-stimulated release in the presence of Ca(2+). It is concluded that capsaicin induces release of substance P from dorsal root ganglion neurons via two mechanisms, one requiring extracellular Ca(2+) and the intact synaptosomal-associated protein 25 kDa (SNAP-25), and the other independent of extracellular Ca(2+) and not involving SNAP-25.

Cervical cancer screening knowledge and practices among Korean-American women.

Cervical cancer is one of the most common cancers of American women. The Papanicolaou (Pap) smear test for cervical screening is a widely used and effective means to reduce the morbidity and mortality rate from cervical cancer through early detection. Despite these benefits, many women have never been screened or are not screened at regular intervals. The purpose of this study was to examine cervical cancer screening knowledge and practices of Korean-American women. The sample consisted of 159 Korean-American women, 40 to 69 years of age. The 1987 Cancer Control Supplement questionnaire was translated into Korean and used to collect data. Twenty-six percent of the respondents never heard of the Pap smear test. Only 34% of respondents reported having had a Pap smear test for screening. The most frequently cited reason for not having had a Pap smear test was absence of disease symptoms. Results indicate that education and usual sources of health care were significant factors related to having heard of or having had a Pap smear test. The findings from this study have important implications for health practitioners and policy makers who serve this ethnic population.

Ionomycin induced changes in intracellular free calcium in SH-SY5Y human neuroblastoma cells: sources of calcium and effects on [3H]-noradrenaline release.

In adherent SH-SY5Y human neuroblastoma cells cultured for 14 days to promote uptake and release of [3H]-noradrenaline, ionomycin induced a biphasic elevation of the intracellular [Ca2+] ([Ca2+]i). This consisted of a rapid transient elevation followed by a marked, persistent secondary elevation. Further study indicated that the peak [Ca2+]i elevation was dependent upon intracellular Ca2+ whilst the secondary elevation was dependent upon extracellular Ca2+. This profile of response and dependence upon intracellular and extracellular sources of Ca2+ was similar to that evoked by the muscarinic agonist, methacholine but was independent of inositol 1,4,5-trisphosphate generation. Ionomycin also stimulated the release of [3H]-noradrenaline from preloaded cells. Both intracellular and extracellular sources of Ca2+ were needed for the full response and synergised to effect release. Thus, in adherent SH-SY5Y cells, ionomycin elevates [Ca2+]i in a complex way in a manner partly analogous to the elevation of [Ca2+]i by agonists of phosphoinositidase C-linked receptors. Furthermore the effects of [Ca2+]i elevation on [3H]-noradrenaline release by these two processes are similar. Such functional consequences may, however, differ under circumstances where the profile and source of Ca2+ for ionomycin-mediated changes differs to that of receptor agonists.

Mobilization of inositol 1,4,5-trisphosphate-sensitive Ca2+ stores supports bradykinin- and muscarinic-evoked release of [3H] noradrenaline from SH-SY5Y cells.

The human neuroblastoma cell line SH-SY5Y, maintained at confluence for 14 days, released [3H]-noradrenaline ([3H]NA) when stimulated with either the muscarinic receptor agonist methacholine or bradykinin. The major fraction of release was rapid, occurring in < 10 s, whereas nicotine-evoked release was slower. When the extracellular [Ca2+]e) was buffered to approximately 50-100 nM, release evoked by nicotine was abolished, whereas that in response to methacholine or bradykinin was reduced by approximately 50% with EC50 values of -5.46 +/- 0.05 M and -7.46 +/- 0.06 M (log 10), respectively. Methacholine and bradykinin also produced rapid elevations of both inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and intracellular free [Ca2+] ([Ca2+]i). These elevations were reduced at low [Ca2+]e and under these conditions the EC50 values for peak elevation of [Ca2+]i were -6.00 +/- 0.14 M for methacholine and -7.95 +/- 0.34 M for bradykinin (n = 3 for all EC50 determinations). At low [Ca2+]e, depletion of nonmitochondrial intracellular Ca2+ stores with the Ca(2+)-ATPase inhibitor thapsigargin produced a transient small elevation of [Ca2+]i and a minor release of [3H]NA. At low [Ca2+]e, thapsigargin abolished elevation of [Ca2+]i in response to methacholine and bradykinin and completely inhibited their stimulation of [3H]NA release. It is proposed, therefore, that Ca2+ release from Ins (1,4,5)P3-sensitive stores is a major trigger of methacholine- and bradykinin-evoked [3H]NA release in SH-SY5Y cells.

[Hemangioma, the most frequent hepatic tumor. Diagnosis with dynamic CAT]. / El hemangioma, tumor hepático más frecuente. Diagnóstico con TAC dinámico.

The sensitivity, specificity and predictive values of dynamic CAT with contrast piston-stroke performed at a single cut are described for the diagnosis of hepatic hemangiomas. We analyzed the correlation between the findings obtained through dynamic CAT and those obtained through echography, PAAF, analytic and clinical study of the patients with suspicion of hepatic hemangioma. The following values were obtained: sensitivity 92.3%; specificity 50%; VPP 88.8%; VPN 60%; and global diagnostic affectivity 84.37%. According to these results, we think that dynamic CAT is a highly reliable test for the diagnosis of hepatic hemangiomas.

Stimulation of phospholipase C in cultured microvascular endothelial cells from human frontal lobe by histamine, endothelin and purinoceptor agonists.

1. Cultures of endothelial cells derived from the microvasculature of human frontal lobe have been investigated for phospholipase C (PLC) responses to histamine, endothelins and purinoceptor agonists. 2. Using cells prelabelled with [3H]-inositol and measuring total [3H]-inositol (poly)phosphates, histamine acting at H1 receptors stimulated a substantial response with an EC50 of about 10 microM. 3. Endothelin-1 also gave a clear stimulation of phosphoinositide-specific phospholipase C. Both concentration-response curves and binding curves showed effective responses and binding in the rank order of endothelin-1 > sarafotoxin S6b > endothelin-3, suggesting an ETA receptor. 4. Assay of total [3H]-inositol (poly)phosphates showed no response to the purinoceptor agonists, 2-methylthioadenosine 5'-trisphosphate (2MeSATP), adenosine 5'-O-(3-thiotrisphosphate) (ATP gamma S) or beta,gamma-methylene ATP. Both ATP and UTP gave a small PLC response. 5. Similarly, when formation of [32P]-phosphatidic acid from cells prelabelled with 32Pi was used as an index of both PLC and phospholipase D, a small response to ATP and UTP was seen but there was no response to the other purinoceptor agonists tested. 6. Study by mass assay of stimulation by ATP of inositol (1,4,5) trisphosphate accumulation revealed a transient response in the first few seconds, a decline to basal, followed by a small sustained response. 7. These results show that human brain endothelial cells in culture are responsive to histamine and endothelins in a manner which may regulate brain capillary permeability. Purines exert a lesser influence.

Differential heterologous and homologous desensitization of two receptors for ATP (P2y purinoceptors and nucleotide receptors) coexisting on endothelial cells.

Bovine aortic endothelial cells in culture contain two coexisting phosphoinositidase C-linked receptors for ATP, the P2y-purinoceptors [for which 2-methylthio-ATP (2MeSATP) is a selective agonist] and the nucleotide (or P2u) receptors (for which UTP is a selective agonist). Here we have investigated the occurrence of homologous and heterologous desensitization of these two receptors and the involvement of protein kinase C-dependent mechanisms. Measuring total [3H]inositol (poly)phosphate accumulation in the presence of lithium, we showed that with long (15-min) stimulations with UTP or 2MeSATP desensitization occurred to a maximum of 40% within several minutes of preexposure to either agonist, i.e., with this procedure there is no difference between the heterologous and the homologous experimental design. In the remainder of the experiments reported we measured inositol-1,4,5-trisphosphate mass levels, using a protocol of 5-min preincubation, 2-min wash, and 5-sec stimulation. We found that preincubation with either agonist led to desensitization of the response to the same agonist of about 40%. However, whereas preincubation with 2MeSATP did not affect the subsequent response to UTP, preincubation with UTP did attenuate the 2MeSATP response. These results demonstrate that homologous desensitization occurs with both P2Y and nucleotide receptors but that heterologous desensitization follows only from activation of the nucleotide receptors. Preincubation with the protein kinase C inhibitor Ro 31-8220 enhanced the subsequent inositol-1,4,5-trisphosphate response to 2MeSATP but did not affect the desensitization of this response by preincubation with the same agonist. However, whereas the response to UTP was not enhanced by preincubation with the protein kinase C inhibitor, the desensitization caused by preincubation with UTP was partially inhibited by Ro 31-8220. These results show that multiple desensitizing events occur during the first few minutes of receptor activation and that these events are different for each of the receptors for ATP.