Application of near infra-red laser light increases current threshold in optic nerve consistent with increased Na+-dependent transport.

Increases in the current threshold occur in optic nerve axons with the application of infra-red laser light, whose mechanism is only partly understood. In isolated rat optic nerve, laser light was applied near the site of electrical stimulation, via a flexible fibre optic. Paired applications of light produced increases in threshold that were reduced on the second application, the response recovering with increasing delays, with a time constant of 24 s. 3-min duration single applications of laser light gave rise to a rapid increase in threshold followed by a fade, whose time-constant was between 40 and 50 s. After-effects were sometimes apparent following the light application, where the resting threshold was reduced. The increase in threshold was partially blocked by 38.6 mM Li+ in combination with 5  µ M bumetanide, a manoeuvre increasing refractoriness and consistent with axonal depolarization. Assessing the effect of laser light on the nerve input resistance ruled out a previously suggested fall in myelin resistance as contributing to threshold changes. These data appear consistent with an axonal membrane potential that partly relies on temperature-dependent electroneutral Na+ influx, and where fade in the response to the laser may be caused by a gradually diminishing Na+ pump-induced hyperpolarization, in response to falling intracellular [Na+].

Changing the firing threshold for normal optic nerve axons by the application of infra-red laser light.

Normal optic nerve axons exhibit a temperature dependence, previously explained by a membrane potential hyperpolarization on warming. We now report that near infra-red laser light, delivered via a fibre optic light guide, also affects axonal membrane potential and threshold, at least partly through a photo-thermal effect. Application of light to optic nerve, at the recording site, gave rise to a local membrane potential hyperpolarization over a period of about a minute, and increased the size of the depolarizing after potential. Application near the site of electrical stimulation reversibly raised current-threshold, and the change in threshold recorded over minutes of irradiation was significantly increased by the application of the Ih blocker, ZD7288 (50 µM), indicating Ih limits the hyperpolarizing effect of light. Light application also had fast effects on nerve behaviour, increasing threshold without appreciable delay (within seconds), probably by a mechanism independent of kinetically fast K+ channels and Na+ channel inactivation, and hypothesized to be caused by reversible changes in myelin function.

The effects of temperature on the biophysical properties of optic nerve F-fibres.

In multiple sclerosis, exacerbation of symptoms with rising body temperature is associated with impulse conduction failure. The mechanism is not fully understood. Remarkably, normal optic nerve axons also show temperature dependent effects, with a fall in excitability with warming. Here we show two properties of optic nerve axons, accommodation and inward rectification (Ih), respond to temperature changes in a manner consistent with a temperature dependent membrane potential. As we could find no evidence for the functional expression of KV7.2 in the axons, using the K+ channel blocker tetraethylammonium ions, we suggest this may explain the membrane potential lability. In order to understand how the axonal membrane potential may show temperature dependence, we have developed a hypothesis involving the electroneutral movement of Na+ ions across the axon membrane, that increases with increasing temperature with an appropriate Q10. Part, but probably not all, of the electroneutral Na+ movement is eliminated by removing extracellular Cl- or exposure to bumetanide, consistent with the involvement of the transporter NKCC1. Numerical simulation suggests a change in membrane potential of - 15-20 mV mimics altering temperature between room and physiological in the largest axons.

Painful and painless mutations of SCN9A and SCN11A voltage-gated sodium channels.

Chronic pain is a global problem affecting up to 20% of the world's population and has a significant economic, social and personal cost to society. Sensory neurons of the dorsal root ganglia (DRG) detect noxious stimuli and transmit this sensory information to regions of the central nervous system (CNS) where activity is perceived as pain. DRG neurons express multiple voltage-gated sodium channels that underlie their excitability. Research over the last 20 years has provided valuable insights into the critical roles that two channels, NaV1.7 and NaV1.9, play in pain signalling in man. Gain of function mutations in NaV1.7 cause painful conditions while loss of function mutations cause complete insensitivity to pain. Only gain of function mutations have been reported for NaV1.9. However, while most NaV1.9 mutations lead to painful conditions, a few are reported to cause insensitivity to pain. The critical roles these channels play in pain along with their low expression in the CNS and heart muscle suggest they are valid targets for novel analgesic drugs.

The dichotomous effects of caffeine on homologous recombination in mammalian cells.

This study was initiated to examine the effects of caffeine on the DNA damage response (DDR) and homologous recombination (HR) in mammalian cells. A 5 mM caffeine treatment caused the cell cycle to stall at G2/M and cells eventually underwent apoptosis. Caffeine exposure also induced a strong DDR along with subsequent activation of wildtype p53 protein. An unexpected observation was the caffeine-induced depletion of Rad51 (and Brca2) proteins. Consequently, caffeine-treated cells were expected to be inefficient in HR. However, a dichotomy in the HR response of cells to caffeine treatment was revealed. Caffeine treatment rendered cells significantly better at performing the nascent DNA synthesis that accompanies the early strand invasion steps of HR. Additionally, caffeine treatment increased chromatin accessibility and elevated the efficiency of illegitimate recombination. Conversely, the increase in nascent DNA synthesis did not translate into a higher number of gene targeting events. Thus, prolonged caffeine exposure stalls the cell cycle, induces a p53-mediated apoptotic response and a down-regulation of critical HR proteins, and for reasons discussed, stimulates early steps of HR, but not the formation of complete recombination products.

Lacosamide efficacy and tolerability in clinical practice - Post marketing analysis from a single dedicated epilepsy center.

OBJECTIVES: Post marketing analysis of anti-epileptic drug (AED) efficacy and tolerability is of great value to the clinician since it is more representative of clinical practice than clinical trial data. We analyzed our experience with lacosamide (LCM) in patients treated after marketing. PATIENTS AND METHODS: We identified all patients who were treated with LCM during the four year period after marketing, excluding patients who were in clinical trials. We recorded demographic data and analyzed efficacy and tolerability in patients who had at least one follow up visit or telephone call 3 months after the initiation of LCM. RESULTS: A total of 165 patients met our inclusion criteria. The mean age was 41 years. The majority of the cohort had focal epilepsy (146 patients) (88.4%). The mean duration of treatment was 31.2 months. Eighty one patients (49.1%) were continuing LCM at last follow up. Adverse effects (AEs) and discontinuation were significantly more common when LCM was added to one or more Na-channel blocking agents (NCB) (p = 0.0003 and 0.17). The 50% responder rate was 26% at 3 months and increased to 49% at 36 months. Patients were more likely to continue the drug and less likely to have AEs with slower titration over >4 weeks (p = 0.02 for each). Four or more previously failed AEDs predicted poorer response rate compared to three or less AEDs (p = 0.001). CONCLUSION: LCM use in clinical practice was associated with greater rate of seizure freedom than in clinical trials. Discontinuation and occurrence of AEs were significantly more likely with faster titration and adding LCM to NCB agents.

Serum Albumin's Protective Inhibition of Amyloid-ß Fiber Formation Is Suppressed by Cholesterol, Fatty Acids and Warfarin.

Central to Alzheimer's disease (AD) pathology is the assembly of monomeric amyloid-ß peptide (Aß) into oligomers and fibers. The most abundant protein in the blood plasma and cerebrospinal fluid is human serum albumin. Albumin can bind to Aß and is capable of inhibiting the fibrillization of Aß at physiological (µM) concentrations. The ability of albumin to bind Aß has recently been exploited in a phase II clinical trial, which showed a reduction in cognitive decline in AD patients undergoing albumin-plasma exchange. Here we explore the equilibrium between Aß monomer, oligomer and fiber in the presence of albumin. Using transmission electron microscopy and thioflavin-T fluorescent dye, we have shown that albumin traps Aß as oligomers, 9 nm in diameter. We show that albumin-trapped Aß oligomeric assemblies are not capable of forming ion channels, which suggests a mechanism by which albumin is protective in Aß-exposed neuronal cells. In vivo albumin binds a variety of endogenous and therapeutic exogenous hydrophobic molecules, including cholesterol, fatty acids and warfarin. We show that these molecules bind to albumin and suppress its ability to inhibit Aß fiber formation. The interplay between Aß, albumin and endogenous hydrophobic molecules impacts Aß assembly; thus, changes in cholesterol and fatty acid levels in vivo may impact Aß fibrillization, by altering the capacity of albumin to bind Aß. These observations are particularly intriguing given that high cholesterol or fatty acid diets are well-established risk factors for late-onset AD.

Big conductance calcium-activated potassium channel openers control spasticity without sedation.

BACKGROUND AND PURPOSE: Our initial aim was to generate cannabinoid agents that control spasticity, occurring as a consequence of multiple sclerosis (MS), whilst avoiding the sedative side effects associated with cannabis. VSN16R was synthesized as an anandamide (endocannabinoid) analogue in an anti-metabolite approach to identify drugs that target spasticity. EXPERIMENTAL APPROACH: Following the initial chemistry, a variety of biochemical, pharmacological and electrophysiological approaches, using isolated cells, tissue-based assays and in vivo animal models, were used to demonstrate the activity, efficacy, pharmacokinetics and mechanism of action of VSN16R. Toxicological and safety studies were performed in animals and humans. KEY RESULTS: VSN16R had nanomolar activity in tissue-based, functional assays and dose-dependently inhibited spasticity in a mouse experimental encephalomyelitis model of MS. This effect occurred with over 1000-fold therapeutic window, without affecting normal muscle tone. Efficacy was achieved at plasma levels that are feasible and safe in humans. VSN16R did not bind to known CB1 /CB2 /GPPR55 cannabinoid-related receptors in receptor-based assays but acted on a vascular cannabinoid target. This was identified as the major neuronal form of the big conductance, calcium-activated potassium (BKCa ) channel. Drug-induced opening of neuronal BKCa channels induced membrane hyperpolarization, limiting excessive neural-excitability and controlling spasticity. CONCLUSIONS AND IMPLICATIONS: We identified the neuronal form of the BKCa channel as the target for VSN16R and demonstrated that its activation alleviates neuronal excitability and spasticity in an experimental model of MS, revealing a novel mechanism to control spasticity. VSN16R is a potential, safe and selective ligand for controlling neural hyper-excitability in spasticity.

Diuretic-sensitive electroneutral Na+ movement and temperature effects on central axons.

KEY POINTS: Optic nerve axons get less excitable with warming. F-fibre latency does not shorten at temperatures above 30°C. Action potential amplitude falls when the Na+ -pump is blocked, an effect speeded by warming. Diuretics reduce the rate of action potential fall in the presence of ouabain. Our data are consistent with electroneutral entry of Na+ occurring in axons and contributing to setting the resting potential. ABSTRACT: Raising the temperature of optic nerve from room temperature to near physiological has effects on the threshold, refractoriness and superexcitability of the shortest latency (fast, F) nerve fibres, consistent with hyperpolarization. The temperature dependence of peak impulse latency was weakened at temperatures above 30°C suggesting a temperature-sensitive process that slows impulse propagation. The amplitude of the supramaximal compound action potential gets larger on warming, whereas in the presence of bumetanide and amiloride (blockers of electroneutral Na+ movement), the action potential amplitude consistently falls. This suggests a warming-induced hyperpolarization that is reduced by blocking electroneutral Na+ movement. In the presence of ouabain, the action potential collapses. This collapse is speeded by warming, and exposure to bumetanide and amiloride slows the temperature-dependent amplitude decline, consistent with a warming-induced increase in electroneutral Na+ entry. Blocking electroneutral Na+ movement is predicted to be useful in the treatment of temperature-dependent symptoms under conditions with reduced safety factor (Uhthoff's phenomenon) and provide a route to neuroprotection.

Ion Channel Formation by Amyloid-ß42 Oligomers but Not Amyloid-ß40 in Cellular Membranes.

A central hallmark of Alzheimer's disease is the presence of extracellular amyloid plaques chiefly consisting of amyloid-ß (Aß) peptides in the brain interstitium. Aß largely exists in two isoforms, 40 and 42 amino acids long, but a large body of evidence points to Aß(1-42) rather than Aß(1-40) as the cytotoxic form. One proposed mechanism by which Aß exerts toxicity is the formation of ion channel pores that disrupt intracellular Ca2+ homeostasis. However, previous studies using membrane mimetics have not identified any notable difference in the channel forming properties between Aß(1-40) and Aß(1-42). Here, we tested whether a more physiological environment, membranes excised from HEK293 cells of neuronal origin, would reveal differences in the relative channel forming ability of monomeric, oligomeric, and fibrillar forms of both Aß(1-40) and Aß(1-42). Aß preparations were characterized with transmission electron microscopy and thioflavin T fluorescence. Aß was then exposed to the extracellular face of excised membranes, and transmembrane currents were monitored using patch clamp. Our data indicated that Aß(1-42) assemblies in oligomeric preparations form voltage-independent, non-selective ion channels. In contrast, Aß(1-40) oligomers, fibers, and monomers did not form channels. Ion channel conductance results suggested that Aß(1-42) oligomers, but not monomers and fibers, formed three distinct pore structures with 1.7-, 2.1-, and 2.4-nm pore diameters. Our findings demonstrate that only Aß(1-42) contains unique structural features that facilitate membrane insertion and channel formation, now aligning ion channel formation with the differential neurotoxic effect of Aß(1-40) and Aß(1-42) in Alzheimer's disease.

P2Y Receptors Sensitize Mouse and Human Colonic Nociceptors.

Activation of visceral nociceptors by inflammatory mediators contributes to visceral hypersensitivity and abdominal pain associated with many gastrointestinal disorders. Purine and pyrimidine nucleotides (e.g., ATP and UTP) are strongly implicated in this process following their release from epithelial cells during mechanical stimulation of the gut, and from immune cells during inflammation. Actions of ATP are mediated through both ionotropic P2X receptors and metabotropic P2Y receptors. P2X receptor activation causes excitation of visceral afferents; however, the impact of P2Y receptor activation on visceral afferents innervating the gut is unclear. Here we investigate the effects of stimulating P2Y receptors in isolated mouse colonic sensory neurons, and visceral nociceptor fibers in mouse and human nerve-gut preparations. Additionally, we investigate the role of Nav1.9 in mediating murine responses. The application of UTP (P2Y2 and P2Y4 agonist) sensitized colonic sensory neurons by increasing action potential firing to current injection and depolarizing the membrane potential. The application of ADP (P2Y1, P2Y12, and P2Y13 agonist) also increased action potential firing, an effect blocked by the selective P2Y1 receptor antagonist MRS2500. UTP or ADP stimulated afferents, including mouse and human visceral nociceptors, in nerve-gut preparations. P2Y1 and P2Y2 transcripts were detected in 80% and 56% of retrogradely labeled colonic neurons, respectively. Nav1.9 transcripts colocalized in 86% of P2Y1-positive and 100% of P2Y2-positive colonic neurons, consistent with reduced afferent fiber responses to UTP and ADP in Na(v)1.9(-/-) mice. These data demonstrate that P2Y receptor activation stimulates mouse and human visceral nociceptors, highlighting P2Y-dependent mechanisms in the generation of visceral pain during gastrointestinal disease.

Ultrasound-Guided Femoral Nerve Blocks.

Pediatric acute femur fractures are a relatively common major orthopedic injury seen in emergency departments. Providing adequate and safe analgesia is essential while patients await definitive management of these fractures. Opioid medications are typically used to treat fracture-associated pain but have well-known adverse effects including respiratory and central nervous system depression, pruritus, nausea, and allergic reactions. Dose titration of opioids in pediatric patients may be difficult and requires frequent nursing and physician reassessments. Regional anesthesia using ultrasound guidance has been proposed as a reliable and safe method to provide pain relief for this population and to decrease reliance on opioid medications. There is a growing body of literature on the utility and safety of ultrasound-guided femoral nerve blocks for pediatric patients in the acute care setting. This review article covers recent literature on point-of-care ultrasound-guided femoral nerve blocks for pediatric patients, with a discussion of the indications, sonographic anatomy, selection of anesthetics, nerve block technique, and complications. This review supplements the expert supervision and practice required to gain competency.

Acute temperature sensitivity in optic nerve axons explained by an electrogenic membrane potential.

Classical work in squid axon reports resting membrane potential is independent of temperature, but our findings suggest that this is not the case for axons in mammalian optic nerve. Refractory period duration changes over 10 times between 37 °C and room temperature, and afterpotential polarity is also acutely temperature sensitive, inconsistent with changes in temperature impacting nerve function only through altered rates of ion channel gating kinetics. Our evidence suggests that the membrane potential is enhanced by warming, an effect reduced by exposure to ouabain. The temperature dependence can be explained if axonal Na(+)/K(+) ATPase continuously expels Na(+) ions that enter axons largely electroneutrally, thereby adding a substantial electrogenic component to the membrane potential. Block of the Na(+) transporter NKCC1 with bumetanide increases refractoriness, like depolarization, indicating that this is a probable route by which Na(+) enters, raising the expectation that the rate of electroneutral Na(+) influx increases with temperature and suggesting a temperature-dependent transmembrane Na(+) cycle that contributes to membrane potential.

Ability of pediatric emergency medicine physicians to identify anatomic landmarks with the assistance of ultrasound prior to lumbar puncture in a simulated obese model.

OBJECTIVES: Lumbar punctures (LPs) are typically performed using anatomical surface landmarks. However, as body mass index increases, identifying surface landmarks becomes more difficult. Ultrasound has been proposed as a tool for identifying these landmarks prior to LP. This study evaluates the effectiveness of a brief training program in ultrasound identification of anatomical landmarks in a simulated obese model prior to completing an LP. METHODS: Pediatric emergency medicine physicians completed a pretest questionnaire on ultrasound familiarity prior to an educational session. Participants utilized ultrasonography without the assistance of palpation on a simulation LP model saving images for review. Participants attempted LP on phantom models with simulated body mass indices of 35 and 40 kg/m. Time to image acquisition and successful aspiration of cerebrospinal fluid from the model were recorded. Two expert sonologists independently reviewed all images for correct landmark identification. RESULTS: Seven of the 19 participants had previous familiarity with ultrasound. The mean time to lumbar image acquisition significantly improved for all individuals from 176 seconds to 100 seconds (P = 0.003). Comfort level with ultrasound improved (P < 0.001) as well as comfort level in performing a lumbar ultrasound (P < 0.001). Adequate images were obtained in 96% of the attempts (55/57). The success rate at performing LP was 95% (54/57). CONCLUSIONS: After a brief education intervention, pediatric emergency medicine physicians with little to no previous training in ultrasound can obtain adequate lumbar anatomic images and successfully perform LP in a simulated obese model. Comfort level with ultrasound significantly improves with a short course in ultrasound fundamentals.

Multiple roles for NaV1.9 in the activation of visceral afferents by noxious inflammatory, mechanical, and human disease-derived stimuli.

Chronic visceral pain affects millions of individuals worldwide and remains poorly understood, with current therapeutic options constrained by gastrointestinal adverse effects. Visceral pain is strongly associated with inflammation and distension of the gut. Here we report that the voltage-gated sodium channel subtype NaV1.9 is expressed in half of gut-projecting rodent dorsal root ganglia sensory neurons. We show that NaV1.9 is required for normal mechanosensation, for direct excitation and for sensitization of mouse colonic afferents by mediators from inflammatory bowel disease tissues, and by noxious inflammatory mediators individually. Excitatory responses to ATP or PGE2 were substantially reduced in NaV1.9(-/-) mice. Deletion of NaV1.9 substantially attenuates excitation and subsequent mechanical hypersensitivity after application of inflammatory soup (IS) (bradykinin, ATP, histamine, PGE2, and 5HT) to visceral nociceptors located in the serosa and mesentery. Responses to mechanical stimulation of mesenteric afferents were also reduced by loss of NaV1.9, and there was a rightward shift in stimulus-response function to ramp colonic distension. By contrast, responses to rapid, high-intensity phasic distension of the colon are initially unaffected; however, run-down of responses to repeat phasic distension were exacerbated in NaV1.9(-/-) afferents. Finally colonic afferent activation by supernatants derived from inflamed human tissue was greatly reduced in NaV1.9(-/-) mice. These results demonstrate that NaV1.9 is required for persistence of responses to intense mechanical stimulation, contributes to inflammatory mechanical hypersensitivity, and is essential for activation by noxious inflammatory mediators, including those from diseased human bowel. These observations indicate that NaV1.9 represents a high-value target for development of visceral analgesics.

Randomized trial comparing two mass casualty triage systems (JumpSTART versus SALT) in a pediatric simulated mass casualty event.

PURPOSE: Several field triage systems have been developed to rapidly sort patients following a mass casualty incident (MCI). JumpSTART (Simple Triage and Rapid Transport) is a pediatric-specific MCI triage system. SALT (Sort, Assess, Lifesaving interventions, Treat/Transport) has been proposed as a new national standard for MCI triage for both adult and pediatric patients, but it has not been tested in a pediatric population. This pilot study hypothesizes that SALT is at least as good as JumpSTART in triage accuracy, speed, and ease of use in a simulated pediatric MCI. METHODS: Paramedics were invited and randomly assigned to either SALT or JumpSTART study groups. Following randomization, subjects viewed a 15-minute PowerPoint lecture on either JumpSTART or SALT. Subjects were provided with a triage algorithm card for reference and were asked to assign triage categories to 10 pediatric patients in a simulated building collapse. The scenario consisted of 4 children in moulage and 6 high-fidelity pediatric simulators. Injuries and triage categories were based on a previously published MCI scenario. One investigator followed each subject to record time and triage assignment. All subjects completed a post-test survey and structured interview following the simulated disaster. RESULTS: Forty-three paramedics were enrolled. Seventeen were assigned to the SALT group with an overall triage accuracy of 66% ±15%, an overtriage mean rate of 22 ± 16%, and an undertriage rate of 10 ± 9%. Twenty-six participants were assigned to the JumpSTART group with an overall accuracy of 66 ± 12%, an overtriage mean of 23 ±16%, and an undertriage rate of 11.2 ± 11%. Ease of use was not statistically different between the two systems (median Likert value of both systems = 2, p = 0.39) Time to triage per patient was statistically faster in the JumpSTART group (SALT = 34 ± 23 seconds, JumpSTART = 26 ± 19 seconds, p = 0.02). Both systems were prone to cognitive and affective error. CONCLUSION: SALT appears to be at least as good as JumpSTART in overall triage accuracy, overtriage, or undertriage rates in a simulated pediatric MCI. Both systems were considered easy to use. However, JumpSTART was 8 seconds faster per patient in time taken to assign triage designations.

Nascent DNA synthesis during homologous recombination is synergistically promoted by the rad51 recombinase and DNA homology.

In this study, we exploited a plasmid-based assay that detects the new DNA synthesis (3' extension) that accompanies Rad51-mediated homology searching and strand invasion steps of homologous recombination to investigate the interplay between Rad51 concentration and homology length. Mouse hybridoma cells that express endogenous levels of Rad51 display an approximate linear increase in the frequency of 3' extension for homology lengths of 500 bp to 2 kb. At values below ∼500 bp, the frequency of 3' extension declines markedly, suggesting that this might represent the minimal efficient processing segment for 3' extension. Overexpression of wild-type Rad51 stimulated the frequency of 3' extension by ∼3-fold for homology lengths <900 bp, but when homology was >2 kb, 3' extension frequency increased by as much as 10-fold. Excess wild-type Rad51 did not increase the average 3' extension tract length. Analysis of cell lines expressing N-terminally FLAG-tagged Rad51 polymerization mutants F86E, A89E, or F86E/A89E established that the 3' extension process requires Rad51 polymerization activity. Mouse hybridoma cells that have reduced Brca2 (Breast cancer susceptibility 2) due to stable expression of small interfering RNA show a significant reduction in 3' extension efficiency; expression of wild-type human BRCA2, but not a BRCA2 variant devoid of BRC repeats 1-8, rescues the 3' extension defect in these cells. Our results suggest that increased Rad51 concentration and homology length interact synergistically to promote 3' extension, presumably as a result of enhanced Brca2-mediated Rad51 polymerization.

Endogenous levels of Rad51 and Brca2 are required for homologous recombination and regulated by homeostatic re-balancing.

Stable expression of Rad51 siRNA was used to generate mouse hybridoma cell lines in which endogenous Rad51 levels were depleted by as much as 60%. Stable Rad51 knockdowns feature reduced homologous recombination responses. The relative ease with which stable Rad51 knockdowns were recovered was surprising, given the embryonic lethality of Rad51 ablation. Interestingly, Rad51-depleted hybridoma cell lines are characterized by reduced levels of p53 protein. Completely unexpected, was the finding that Rad51-depleted hybridoma cell lines are also reduced for the breast cancer susceptibility 2 (Brca2) protein. Additionally, hybridoma cell lines that are siRNA depleted for mouse Brca2 show a corresponding reduction in Rad51 and p53 proteins. Furthermore, cellular levels of Rad51, Brca2 and p53 can be elevated in these cell lines by ectopic expression of wild-type human Rad51 and wild-type human BRCA2. In marked contrast, hybridoma cell lines that are siRNA depleted for mouse p53 feature relatively normal Rad51 and Brca2 levels. These results suggest that cellular levels of Brca2 and Rad51 are mutually dependent on each other, and that low levels of these proteins provide selective pressure for reduction of p53, which permits cell growth.