Advances in quality control: mouse embryo morphokinetics are sensitive markers of in vitro stress.

STUDY QUESTION: Can time-lapse analysis of cell division timings [morphokinetics (MK)] in mouse embryos detect toxins at concentrations that do not affect blastocyst formation? SUMMARY ANSWER: An MK algorithm enhances assay sensitivity while providing results 24-48 h sooner than the traditional mouse embryo assay (MEA). WHAT IS KNOWN ALREADY: Current quality control testing methodology is sensitive but further improvements are needed to assure optimal culture conditions. MKs of embryo development may detect small variations in culture conditions. STUDY DESIGN: Cross sectional-control versus treatment. Mouse embryo development kinetics of 466 embryos were analyzed according to exposure to various concentrations of toxins and toxic mineral oil. MATERIALS, SETTING, METHODS: Cryopreserved 1-cell embryos from F1 hybrid mice were cultured with cumene hydroperoxide (CH) (0, 2, 4, 6 and 8 µM) and Triton X-100 (TX-100; 0, 0.0008, 0.0012, 0.0016 and 0.002%). Using the Embryoscope, time-lapse images were obtained every 20 min for 120 h in seven focal planes. End-points were timing and pattern of cell division and embryo development. The blastocyst rate (BR) was defined as the percentage of embryos that developed to the expanded blastocyst stage within 96 h. MAIN RESULTS AND THE ROLE OF CHANCE: BR was not affected for embryos cultured in the three lowest concentrations of CH and the four lowest concentrations of TX-100. In contrast, a unique MK model detected all concentrations tested (P < 0.05). The MK model identified toxicity in two lots of toxic mineral oil that did not affect BR (P < 0.05). LIMITATIONS, REASONS FOR CAUTION: A limited number of toxins were used so that the results may not apply to all potential embryo toxins. A larger sample size may also demonstrate other statistically significant developmental kinetic parameters. WIDER IMPLICATIONS OF THE FINDINGS: MKs in mouse embryos are a sensitive and efficient method for quality control testing of in vitro culture conditions. BR, the end-point of traditional quality control assays, did not detect sublethal concentrations of toxins in the culture milieu in our study. This study demonstrates that temporal variation at key developmental stages reflects the quality of the culture environment. An MEA that incorporates MK will provide enhanced sensitivity and faster turn-around times.

Anxiogenic effects of CGRP within the BNST may be mediated by CRF acting at BNST CRFR1 receptors.

Calcitonin gene-related peptide (CGRP) acting within the bed nucleus of the stria terminalis (BNST) increases anxiety as well as neural activation in anxiety-related structures, and mediates behavioral stress responses. Similar effects have been described following intra-ventricular as well as intra-BNST infusions of the stress-responsive neuropeptide, corticotropin releasing factor (CRF). Interestingly, CGRP-positive terminals within the lateral division of the BNST form perisomatic baskets around neurons that express CRF, suggesting that BNST CGRP could exert its anxiogenic effects by increasing release of CRF from these neurons. With this in mind, the present set of experiments was designed to examine the role of CRFR1 signaling in the anxiogenic effects of CGRP within the BNST and to determine whether CRF from BNST neurons contributes to these effects. Consistent with previous studies, we found that 400 ng CGRP infused bilaterally into the BNST increased the acoustic startle response and induced anxiety-like behavior in the elevated plus maze compared to vehicle. Both of these effects were attenuated by 10mg/kg PO of the CRFR1 antagonist, GSK876008. GSK876008 alone did not affect startle. An intra-BNST infusion of the CRFR1 antagonist CP376395 (2 µg) also blocked increases in acoustic startle induced by intra-BNST infusion of CGRP, as did virally-mediated siRNA knockdown of CRF expression locally within the BNST. Together, these results suggest that the anxiogenic effects of intra-BNST CGRP may be mediated by CRF from BNST neurons acting at local CRFR1 receptors.

Effects of continuously enhanced corticotropin releasing factor expression within the bed nucleus of the stria terminalis on conditioned and unconditioned anxiety.

The lateral division of the bed nucleus of the stria terminalis (BNST), which forms part of the circuitry regulating fear and anxiety, contains a large number of neurons expressing corticotropin releasing factor (CRF), a neuropeptide that has a prominent role in the etiology of fear- and anxiety-related psychopathologies. Stress increases CRF expression within BNST neurons, implicating these cells in stress- and anxiety-related behaviors. These experiments examined the effect of chronically enhanced CRF expression within BNST neurons on conditioned and unconditioned anxiety-related behavior by using a lentiviral vector containing a promoter that targets CRF gene overexpression (OE) to CRFergic cells. We found that BNST CRF-OE did not affect unconditioned anxiety-like responses in the elevated plus maze or basal acoustic startle amplitude. CRF-OE induced before training weakened sustained fear (conditioned anxiety); when induced after conditioning, CRF-OE increased expression of the conditioned emotional memory. Increased BNST CRF expression did not affect plasma corticosterone concentration but did decrease CRFR1 receptor density within the BNST and CRFR2 receptor density within the dorsal portion of the caudal dorsal raphe nucleus. These data raise the possibility that the observed behavioral effects may be mediated by enhanced CRF receptor signaling or compensatory changes in CRF receptor density within these structures. Together, these studies demonstrate that CRF neurons within the lateral BNST modulate conditioned anxiety-like behaviors and also suggest that enhanced CRF expression within these neurons may contribute to inappropriate regulation of emotional memories.

Selective participation of the bed nucleus of the stria terminalis and CRF in sustained anxiety-like versus phasic fear-like responses.

The medial division of the central nucleus of the amygdala (CeA(M)) and the lateral division of the bed nucleus of the stria terminalis (BNST(L)) are closely related. Both receive projections from the basolateral amygdala (BLA) and both project to brain areas that mediate fear-influenced behaviors. In contrast to CeA(M) however, initial attempts to implicate the BNST in conditioned fear responses were largely unsuccessful. More recent studies have shown that the BNST does participate in some types of anxiety and stress responses. Here, we review evidence suggesting that the CeA(M) and BNST(L) are functionally complementary, with CeA(M) mediating short- but not long-duration threat responses (i.e., phasic fear) and BNST(L) mediating long- but not short-duration responses (sustained fear or 'anxiety'). We also review findings implicating the stress-related peptide corticotropin-releasing factor (CRF) in sustained but not phasic threat responses, and attempt to integrate these findings into a neural circuit model which accounts for these and related observations.

A novel mouse model of X-linked cardiac hypertrophy.

We recovered a novel mouse mutant exhibiting neonatal lethality associated with severe fetal cardiac hypertrophy and with some adult mice dying suddenly with left ventricular hypertrophic cardiomyopathy. Using Doppler echocardiography, we screened surviving adult mice in this mutant line for cardiac hypertrophy. Cardiac dimensions were obtained either from two-dimensional images collected using a novel ECG-gated ultra-high-frequency ultrasound system or by traditional M-mode imaging on a clinical ultrasound system. These analyses identified, among the littermates, two populations of mice: those with apparent cardiac hypertrophy with hypercontractile function characterized by ejection fraction of 75-80%, and normal littermates with ejection fraction of 53-55%. Analysis of the ECG-gated two-dimensional cines indicated that the hypertrophy was of the nonobstructive type. Further analysis of heart-to-body weight ratio confirmed the ultrasound diagnosis of left ventricular hypertrophic cardiomyopathy. Histopathology showed increased ventricular wall thickness, enlarged myocyte size, and mild myofiber disarray. Ultrastructural analysis by electron microscopy revealed mitochondria hyperproliferation and dilated sarcoplasmic reticulum. Genome scanning using microsatellite DNA markers mapped the mutation to the X chromosome. DNA sequencing showed no mutations in the coding regions of several candidate genes on the X chromosome, including several known to be associated with left ventricular hypertrophic cardiomyopathy. These findings suggest that this mouse line may harbor a mutation in a novel gene causing X-linked cardiomyopathy.

Expression of genes regulating chromosome segregation, the cell cycle and apoptosis during human preimplantation development.

BACKGROUND: Appropriate gene expression is vital for the regulation of developmental processes. Despite this fact there is a remarkable paucity of information concerning gene activity during preimplantation development. METHODS: We employed reverse transcription and real-time fluorescent PCR to quantify the expression of nine genes (BRCA1, BRCA2, ATM, TP53, RB1, MAD2, BUB1, APC and beta-actin) in oocytes and embryos. A full characterization of all genes was achieved in 42 embryos and four oocytes. The genes analysed have a variety of important cellular functions. RESULTS: Oocytes displayed relatively high levels of mRNA transcripts, while 2-3-cell embryos were seen to contain very little mRNA from any of the genes examined. Recovery of expression levels was not seen until the 4-cell stage or later, with the presumptive activation of the embryonic genome. Some genes displayed sharp increases in expression in embryos composed of 4-8 cells, but, for most, maximum expression was not achieved until the blastocyst stage. CONCLUSIONS: Our data show that it is possible to define characteristic gene expression profiles for each stage of human preimplantation development. The identification of genes active at defined preimplantation phases may provide clues to the cellular pathways utilized at specific stages of development. Expression of genes that function in DNA repair pathways indicate that DNA damage may be common at the cleavage stage. We suggest that specific patterns of gene expression may be indicative of embryo implantation potential.

Mitogen-activated protein kinase cascade in the basolateral nucleus of amygdala is involved in extinction of fear-potentiated startle.

Previous results indicate that intra-amygdala infusions of NMDA receptor antagonists block the extinction of conditioned fear. Mitogen-activated protein kinase (MAPK) can be activated by NMDA receptor stimulation and is involved in excitatory fear conditioning. Here, we evaluate the role of MAPK within the basolateral amygdala in the extinction of conditioned fear. Rats received 10 light-shock pairings. After 24 hr, fear was assessed by eliciting the acoustic startle reflex in the presence of the conditioned stimulus (CS) (CS-noise trials) and also in its absence (noise-alone trials). Rats subsequently received an intra-amygdala or intrahippocampal infusion of either 20% DMSO or the MAPK inhibitor PD98059 (500 ng/side) followed 10 min later by 30 presentations of the light CS without shock (extinction training). After 24 hr, they were again tested for fear-potentiated startle. PD98059 infusions into the basolateral amygdala but not the hippocampus significantly reduced extinction, which was otherwise evident in DMSO-infused rats. Control experiments indicated that the effect of intra-amygdala PD98059 could not be attributed to lasting damage to the amygdala or to state dependency. These results suggest that a MAPK-dependent signaling cascade within or very near the basolateral amygdala plays an important role in the extinction of conditioned fear.

Physician compensation: rewarding productivity of the knowledge worker.

Designing a physician incentive compensation plan that aligns the demands of managed care with the perceived fairness of income distribution is a key challenge for medical practices today. Rather than focus on traditional productivity measures, managed care requires physicians to demonstrate efficient practice of medicine. Physicians still need to be highly productive; however, they are now required to demonstrate efficiency related to clinical resource management, patient access and service, and evidence-based outcomes. Approaches to the development of physician incentive compensation plans and case examples are offered to assist practices that are transitioning physician compensation from volume-based to efficiency-based indicators.

Involvement of NMDA receptors within the amygdala in short- versus long-term memory for fear conditioning as assessed with fear-potentiated startle.

Pretraining intra-amygdala infusions of the NMDA receptor antagonist. D,L-AP5, block fear-potentiated startle in rats tested 24+ hr after training. This may reflect a failure of either acquisition or retention. To evaluate these alternatives, rats were tested for fear-potentiated startle during fear conditioning (30 light-shock pairings [0.6 mA shock]), as well as 1-30 min and 48 hr after fear conditioning. Amygdala lesions abolished fear-potentiated startle at all train-test intervals. Intra-amygdala AP5 infusions (25 nmol/side) abolished fear-potentiated startle during the long-term test and had partial effects at shorter train-test intervals. When the level of fear-potentiated startle during the short-term test was lowered to that of the 48-hr test (i.e., by training rats with a lower, 0.3 mA footshock), AP5 abolished fear-potentiated startle at each timepoint. Thus, amygdala NMDA receptors appear to participate in the initial acquisition of fear memories.

Skeletor, a novel chromosomal protein that redistributes during mitosis provides evidence for the formation of a spindle matrix.

A spindle matrix has been proposed to help organize and stabilize the microtubule spindle during mitosis, though molecular evidence corroborating its existence has been elusive. In Drosophila, we have cloned and characterized a novel nuclear protein, skeletor, that we propose is part of a macromolecular complex forming such a spindle matrix. Skeletor antibody staining shows that skeletor is associated with the chromosomes at interphase, but redistributes into a true fusiform spindle structure at prophase, which precedes microtubule spindle formation. During metaphase, the spindle, defined by skeletor antibody labeling, and the microtubule spindles are coaligned. We find that the skeletor-defined spindle maintains its fusiform spindle structure from end to end across the metaphase plate during anaphase when the chromosomes segregate. Consequently, the properties of the skeletor-defined spindle make it an ideal substrate for providing structural support stabilizing microtubules and counterbalancing force production. Furthermore, skeletor metaphase spindles persist in the absence of microtubule spindles, strongly implying that the existence of the skeletor-defined spindle does not require polymerized microtubules. Thus, the identification and characterization of skeletor represents the first direct molecular evidence for the existence of a complete spindle matrix that forms within the nucleus before microtubule spindle formation.

Measuring resource utilization in medical group practice.

The staff seems so busy every time you walk by the front office or pass through the nurses' station. The office seems to brim with patients whenever you stop by to chat with the medical director. "Seems" is the operative word--analyzing reality versus appearance may lead to some important insights about the utilization of resources in your practice. This article serves to define and analyze resource utilization in a medical practice by considering examples of staff and facility utilization.

Murine pharmacokinetics of 6-aminonicotinamide (NSC 21206), a novel biochemical modulating agent.

The pyridine nucleotide 6-aminonicotinamide (6AN) was shown recently to sensitize a number of human tumor cell lines to cisplatin in vitro. The present studies were undertaken to compare the drug concentrations and length of exposure required for this sensitization in vitro with the drug exposure that could be achieved in mice in vivo. Human K562 leukemia cells and A549 lung cancer cells were incubated with 6AN for various lengths of time, exposed to cisplatin for 1-2 hr, and assayed for Pt-DNA adducts as well as the ability to form colonies. K562 cells displayed progressive increases in Pt-DNA adducts and cisplatin sensitivity during the first 10 hr of 6AN exposure. An 18-hr 6AN exposure was likewise more effective than a 6-hr 6AN exposure in sensitizing A549 cells to cisplatin. HPLC analysis of 6AN and its metabolite, 6-amino-NAD+, permitted assessment of exposures achieved in vivo after i.v. administration of 10 mg/kg of 6AN to CD2F1 mice. 6AN reached peak serum concentrations of 80-90 microM and was cleared rapidly, with T1/2alpha and T1/2beta values of 7.4 and 31.3 min, respectively. Bioavailability was 80-100% with identical plasma pharmacokinetics after i.p. administration. At least 25% of the 6AN was excreted unchanged in the urine. The metabolite 6-amino-NAD+ was detected in perchloric acid extracts of brain, liver, kidney, and spleen, but not in serum. Efforts to prolong systemic 6AN exposure by administering multiple i.p. doses or using osmotic pumps resulted in lethal toxicity. These results demonstrated that 6AN exposures required to sensitize tumor cells to cisplatin in vitro are difficult to achieve in vivo.

JIL-1: a novel chromosomal tandem kinase implicated in transcriptional regulation in Drosophila.

We have cloned and characterized JIL-1, a novel tandem kinase in Drosophila that associates with the chromosomes throughout the cell cycle. Antibody staining and live imaging of JIL-1-GFP transgenic flies show that JIL-1 localizes to the gene-rich interband regions of larval polytene chromosomes and is upregulated almost 2-fold on the hypertranscribed male X chromosome compared to autosomes. Phylogenetic analysis suggests that JIL-1 together with human MSKs defines a separate family of tandem kinases. That JIL-1 is a functional kinase was demonstrated by autophosphorylation and phosphorylation of histone H3 in vitro. Based on these findings, we propose that JIL-1 may play a role in transcriptional control potentially by regulating chromatin structure.

Phase I trial of dolastatin-10 (NSC 376128) in patients with advanced solid tumors.

Dolastatin-10 (dola-10) is a potent antimitotic peptide, isolated from the marine mollusk Dolabela auricularia, that inhibits tubulin polymerization. Preclinical studies of dola-10 have demonstrated activity against a variety of murine and human tumors in cell cultures and mice models. The purpose of this Phase I clinical trial was to characterize the maximum tolerated dose, pharmacokinetics, and biological effects of dola-10 in patients with advanced solid tumors. Escalating doses of dola-10 were administered as an i.v. bolus every 21 days, using a modified Fibonacci dose escalation schema. Pharmacokinetic studies were performed with the first treatment cycle. Neurological testing was performed on each patient prior to treatment with dola-10, at 6 weeks and at study termination. Thirty eligible patients received a total of 94 cycles (median, 2 cycles; maximum, 14 cycles) of dola-10 at doses ranging from 65 to 455 microg/m2. Dose-limiting toxicity of granulocytopenia was seen at 455 microg/m2 for minimally pretreated patients (two or fewer prior chemotherapy regimens) and 325 microg/m2 for heavily pretreated patients (more than two prior chemotherapy regimens). Nonhematological toxicity was generally mild. Local irritation at the drug injection site was mild and not dose dependent. Nine patients developed new or increased symptoms of mild peripheral sensory neuropathy that was not dose limiting. This toxicity was more frequent in patients with preexisting peripheral neuropathies. Pharmacokinetic studies demonstrated a rapid drug distribution with a prolonged plasma elimination phase (t 1/2z = 320 min). The area under the concentration-time curve increased in proportion to administered dose, whereas the clearance remained constant over the doses studied. Correlation analysis demonstrated a strong relationship between dola-10 area under the concentration-time curve values and decrease from baseline for leukocyte counts. In conclusion, dola-10 administered every 3 weeks as a peripheral i.v. bolus is well tolerated with dose-limiting toxicity of granulocytopenia. The maximum tolerated dose (and recommended Phase II starting dose) is 400 microg/m2 for patients with minimal prior treatment (two or fewer prior chemotherapy regimens) and 325 microg/m2 for patients who are heavily pretreated (more than two prior chemotherapy regimens).

Molecular cloning and characterization of LKv1, a novel voltage-gated potassium channel in leech.

We have cloned a novel voltage-gated K channel, LKv1, in two species of leech. The properties of LKv1 expressed in transiently transfected HEK293 cells is that of a delayed rectifier current. LKv1 may be a major modulator of excitability in leech neurons, since antibody localization studies show that LKv1 is expressed in the soma and axons of all neurons in both the central and peripheral nervous systems. Comparison of the biophysical and pharmacological properties of LKv1 with native voltage-gated conductances in leech neurons suggests that LKv1 may correspond to the previously characterized delayed rectifier current, I(K). Phylogenetic analysis of LKv1 shows that it is related to the Shaker subfamily of voltage-gated K channels although it occupies a separate branch from that of the monophyletic Shaker clade composed of the flatworm, Aplysia, Drosophila, and mammalian Shaker homologs as well as from that of two recently identified Shaker-related K channels in jellyfish. Thus, this analysis indicates that this group of voltage-gated K channels contains several evolutionarily divergent lineages.

Chromatin structure and nuclear remodeling.

Nuclear architecture is remodeled during interphase in response to changes in gene activity as well as to changing structural and functional requirements during cell division. Using the monoclonal antibody mAb2A, we have identified two proteins that appear to play important roles in these processes: JIL-1 is a tandem serine-threonine kinase implicated in the regulation of chromatin structure, whereas Skeletor is a novel protein participating in structural nuclear remodeling during the cell cycle. Antibody staining and live imaging of JIL-1-GFP transgenic flies show that JIL-1 localizes to the gene-rich interband regions of larval polytene chromosomes and is upregulated almost twofold on the hypertranscribed male X chromosome compared with autosomes. We propose that JIL-1 may play a role in transcriptional control potentially by regulating chromatin structure. The other mAb2A antigen, Skeletor, is distributed in a nuclear meshwork pattern that can be observed in stereo pair images to reorganize during the cell cycle to form a spindle-like structure at prometaphase that is distinct from the microtubule spindle apparatus. Taking advantage of the powerful molecular and genetic approaches offered in Drosophila, the study of these two proteins promises to yield new insight into what defines nuclear architecture at the molecular level and how its remodeling is regulated.

6-Aminonicotinamide sensitizes human tumor cell lines to cisplatin.

The nicotinamide analogue 6-aminonicotinamide (6AN) is presently undergoing evaluation as a potential modulator of the action of various antineoplastic treatments. Most previous studies of this agent have focused on a three-drug regimen of chemical modulators that includes 6AN. In the present study, the effect of single-agent 6AN on the efficacy of selected antineoplastic drugs was assessed in vitro. Colony-forming assays using human tumor cell lines demonstrated that pretreatment with 30-250 microM 6AN for 18 h resulted in increased sensitivity to the DNA cross-linking agent cisplatin, with 6-, 11-, and 17-fold decreases in the cisplatin dose that diminishes colony formation by 90% being observed in K562 leukemia cells, A549 non-small cell lung cancer cells, and T98G glioblastoma cells, respectively. Morphological examination revealed increased numbers of apoptotic cells after treatment with 6AN and cisplatin compared to cisplatin alone. 6AN also sensitized cells to melphalan and nitrogen mustard but not to chlorambucil, 4-hydroperoxycyclophosphamide, etoposide, or daunorubicin. In additional studies undertaken to elucidate the mechanism underlying the sensitization to cisplatin, atomic absorption spectroscopy revealed that 6AN had no effect on the rate of removal of platinum (Pt) adducts from DNA. Instead, 6AN treatment was accompanied by an increase in Pt-DNA adducts that paralleled the degree of sensitization. This effect was not attributable to 6AN-induced decreases in glutathione or NAD+, because other agents that depleted these detoxification cofactors (buthionine sulfoximine and 3-acetylpyridine, respectively) did not increase Pt-DNA adducts. On the contrary, 6AN treatment increased cellular accumulation of cisplatin. Further experiments revealed that 6AN was metabolized to 6-aminonicotinamide adenine dinucleotide (6ANAD+). Concurrent administration of nicotinamide and 6AN had minimal effect on cellular 6AN accumulation but abolished the formation of 6ANAD+, the increase in Pt-DNA adducts, and the sensitizing effect of 6AN in clonogenic assays. These observations identify 6AN as a potential modulator of cisplatin sensitivity and suggest that the 6AN metabolite 6ANAD+ exerts this effect by increasing cisplatin accumulation and subsequent formation of Pt-DNA adducts.

Temporal relationships between oxytocin and 13,14-dihydro-15-keto-prostaglandin F2 alpha pulses in ovariectomized ewes.

The primary objective was to evaluate the role of non-ovarian oxytocin in the initiation of pulses of PGF2 alpha, as measured by peripheral concentrations of 13,14-dihydro-15-keto-prostaglandin F2 alpha (PGFM). A 2 x 2 factorial arrangement of estradiol and progesterone treatments was administered to groups of five ewes after ovariectomy on Day 12. Progesterone (10 mg) was administered at 0700 and 1900 hr on Day 12, and then either progesterone or its vehicle was administered on Days 13 and 14. Silastic implants, either empty or containing estradiol, was administered at ovariectomy. Oxytocin and PGFM were measured in jugular blood samples withdrawn from an indwelling catheter at 5-min intervals for 8 hr on Day 15. Statistically significant pulses of oxytocin, presumably of posterior pituitary origin, were detected in all ewes. Approximately one-half of the oxytocin pulses preceded a pulse in PGFM concentrations by 10 min or less. These pulses tended (P = 0.09) to have a longer duration than those not linked to pulses of PGFM. The number of PGFM pulses that followed or did not follow an oxytocin pulse by 10 min or less was similar (P > 0.2). The amplitude and duration of oxytocin-linked PGFM pulses were greater (P = 0.05) than non-linked pulses. Although several explanations for the lower than anticipated temporal relationship between oxytocin and PGFM pulses are possible, the finding that oxytocin-related PGFM pulses are distinguishable from other pulses is consistent with the concept that oxytocin initiates robust pulses in PGF2 alpha secretion.

Double dissociation between the involvement of the bed nucleus of the stria terminalis and the central nucleus of the amygdala in startle increases produced by conditioned versus unconditioned fear.

The amplitude of the acoustic startle response is reliably enhanced when elicited in the presence of bright light (light-enhanced startle) or in the presence of cues previously paired with shock (fear-potentiated startle). Light-enhanced startle appears to reflect an unconditioned response to an anxiogenic stimulus, whereas fear-potentiated startle reflects a conditioned response to a fear-eliciting stimulus. We examine the involvement of the basolateral nucleus of the amygdala, the central nucleus of the amygdala, and the bed nucleus of the stria terminalis in both phenomena. Immediately before light-enhanced or fear-potentiated startle testing, rats received intracranial infusions of the AMPA receptor antagonist 2, 3-dihydroxy-6-nitro-7-sulphamoylbenzo(F)-quinoxaline (3 microg) or PBS. Infusions into the central nucleus of the amygdala blocked fear-potentiated but not light-enhanced startle, and infusions into the bed nucleus of the stria terminalis blocked light-enhanced but not fear-potentiated startle. Infusions into the basolateral amygdala disrupted both phenomena. These findings indicate that the neuroanatomical substrates of fear-potentiated and light-enhanced startle, and perhaps more generally of conditioned and unconditioned fear, may be anatomically dissociated.

Anxiogenic effects of high illumination levels assessed with the acoustic startle response in rats.

Previous studies have shown that the amplitude of the acoustic startle reflex is increased by the presentation of aversive stimuli. In the present study, the amplitude of acoustic startle in rats was increased by exposure to high illumination levels. The effect was directly related to the intensity (0, 8, 70, and 700 footlamberts) of illumination (experiment I); was blocked by the anxiolytic compound buspirone (experiment II); and showed little or no habituation with repeated testing (experiment III). These results suggest that the elevation of startle amplitude by light may reflect an unconditioned anxiogenic effect of high illumination levels. The possible utility of this phenomenon as an animal model of anxiety is discussed.