Nef defect attenuates HIV viremia and immune dysregulation in the bone marrow-liver-thymus-spleen (BLTS) humanized mouse model.

In vitro studies have shown that deletion of nef and deleterious mutation in the Nef dimerization interface attenuates HIV replication and associated pathogenesis. Humanized rodents with human immune cells and lymphoid tissues are robust in vivo models for investigating the interactions between HIV and the human immune system. Here, we demonstrate that nef deletion impairs HIV replication and HIV-induced immune dysregulation in the blood and human secondary lymphoid tissue (human spleen) in bone marrow-liver-thymus-spleen (BLTS) humanized mice. Furthermore, we also show that nef defects (via deleterious mutations in the dimerization interface) impair HIV replication and HIV-induced immune dysregulation in the blood and human spleen in BLTS-humanized mice. We demonstrate that the reduced replication of nef-deleted and nef-defective HIV is associated with robust antiviral innate immune response, and T helper 1 response. Our results support the proposition that Nef may be a therapeutic target for adjuvants in HIV cure strategies.

Molecular signatures of normal pressure hydrocephalus: a large-scale proteomic analysis of cerebrospinal fluid.

Given the persistent challenge of differentiating idiopathic Normal Pressure Hydrocephalus (iNPH) from similar clinical entities, we conducted an in-depth proteomic study of cerebrospinal fluid (CSF) in 28 shunt-responsive iNPH patients, 38 Mild Cognitive Impairment (MCI) due to Alzheimer's disease, and 49 healthy controls. Utilizing the Olink Explore 3072 panel, we identified distinct proteomic profiles in iNPH that highlight significant downregulation of synaptic markers and cell-cell adhesion proteins. Alongside vimentin and inflammatory markers upregulation, these results suggest ependymal layer and transependymal flow dysfunction. Moreover, downregulation of multiple proteins associated with congenital hydrocephalus (e.g., L1CAM, PCDH9, ISLR2, ADAMTSL2, and B4GAT1) points to a possible shared molecular foundation between congenital hydrocephalus and iNPH. Through orthogonal partial least squares discriminant analysis (OPLS-DA), a panel comprising 13 proteins has been identified as potential diagnostic biomarkers of iNPH, pending external validation. These findings offer novel insights into the pathophysiology of iNPH, with implications for improved diagnosis.

Assessment of Preanalytical Cerebrospinal Fluid Handling and Storage Factors on Measurement of Aß1-42, Aß1-40, and pTau181 Using an Automated Chemiluminescent Platform.

BACKGROUND: Standardizing cerebrospinal fluid (CSF) laboratory protocols will improve the reliability and availability of clinical biomarker testing required for prescription of novel Alzheimer disease (AD) therapies. This study evaluated several preanalytical handling and storage factors common to ß-amyloid1-42 (Aß1-42), ß-amyloid1-40 (Aß1-40), and phosphorylated tau (pTau181) concentrations including storage at different temperatures, extended cap contact, various mixing methods, and multiple freeze-thaw cycles. METHODS: Aß1-42, Aß1-40, and pTau181 concentrations were measured using LUMIPULSE G1200 automated assays. Samples were collected in polypropylene tubes of various volumes. Sample cap-contact was evaluated by storing samples in upright and inverted positions at either 4°C for 1 week or -80°C for 1 month. To assess mixing methods, samples were freeze-thawed and mixed by inversion, vortex, horizontal roller, or unmixed prior to assay sampling. The impact of successive freeze-thaw cycles was assessed through freezing, thawing, and analyzing CSF samples. RESULTS: Short-term storage at 4°C did not affect Aß1-42, Aß1-40, or pTau181 measurements in any tube type. Tube cap contact affected Aß1-42 in 2.5 mL tubes and pTau181 levels in 10 mL tubes. No difference was observed between mixing methods. After 4 freeze-thaw cycles, Aß1-42 significantly decreased but Aß1-40 remained unchanged. Utilizing the Aß1-42/Aß1-40 ratio, Aß1-42 values normalized, maintaining ratio values within ±5% of baseline measurements. CONCLUSIONS: Storage of CSF at 4°C for 1 week or -80°C for 1 month did not significantly affect Aß1-42, Aß1-40, pTau181, or associated ratio measurements. Tube cap-contact impacted pTau181 and pTau181/Aß1-42 values in larger tubes. Mixing methods are equivalent. The Aß1-42/Aß1-40 ratio compensates for freeze-thaw variability up to 4 cycles.

Molecular Signatures of Normal Pressure Hydrocephalus: A Largescale Proteomic Analysis of Cerebrospinal Fluid.

Given the persistent challenge of differentiating idiopathic Normal Pressure Hydrocephalus (iNPH) from similar clinical entities, we conducted an in-depth proteomic study of cerebrospinal fluid (CSF) in 28 shunt-responsive iNPH patients, 38 Mild Cognitive Impairment (MCI) due to Alzheimer's disease, and 49 healthy controls. Utilizing the Olink Explore 3072 panel, we identified distinct proteomic profiles in iNPH that highlight significant downregulation of synaptic markers and cell-cell adhesion proteins. Alongside vimentin and inflammatory markers upregulation, these results suggest ependymal layer and transependymal flow dysfunction. Moreover, downregulation of multiple proteins associated with congenital hydrocephalus (e.g., L1CAM, PCDH9, ISLR2, ADAMTSL2, and B4GAT1) points to a possible shared molecular foundation between congenital hydrocephalus and iNPH. Through orthogonal partial least squares discriminant analysis (OPLS-DA), a panel comprising 13 proteins has been identified as potential diagnostic biomarkers of iNPH, pending external validation. These findings offer novel insights into the pathophysiology of iNPH, with implications for improved diagnosis.

Exploratory Assessment of Proteomic Network Changes in Cerebrospinal Fluid of Mild Cognitive Impairment Patients: A Pilot Study.

(1) Background: Despite the existence of well-established, CSF-based biomarkers such as amyloid-ß and phosphorylated-tau, the pathways involved in the pathophysiology of Alzheimer's disease (AD) remain an active area of research. (2) Methods: We measured 3072 proteins in CSF samples of AD-biomarker positive mild cognitive impairment (MCI) participants (n = 38) and controls (n = 48), using the Explore panel of the Olink proximity extension assay (PEA). We performed group comparisons, association studies with diagnosis, age, and APOE ε4 status, overrepresentation analysis (ORA), and gene set enrichment analysis (GSEA) to determine differentially expressed proteins and dysregulated pathways. (3) Results: GSEA results demonstrated an enrichment of granulocyte-related and chemotactic pathways (core enrichment proteins: ITGB2, ITGAM, ICAM1, SELL, SELP, C5, IL1A). Moreover, some of the well-replicated, differentially expressed proteins in CSF included: ITGAM, ITGB2, C1QA, TREM2, GFAP, NEFL, MMP-10, and a novel tau-related marker, SCRN1. (4) Conclusion: Our results highlight the upregulation of neuroinflammatory pathways, especially chemotactic and granulocyte recruitment in CSF of early AD patients.

Delayed skeletal development and IGF-1 deficiency in a mouse model of lysinuric protein intolerance.

SLC7A7 deficiency, or lysinuric protein intolerance (LPI), causes loss of function of the y+LAT1 transporter critical for efflux of arginine, lysine and ornithine in certain cells. LPI is characterized by urea cycle dysfunction, renal disease, immune dysregulation, growth failure, delayed bone age and osteoporosis. We previously reported that Slc7a7 knockout mice (C57BL/6×129/SvEv F2) recapitulate LPI phenotypes, including growth failure. Our main objective in this study was to characterize the skeletal phenotype in these mice. Compared to wild-type littermates, juvenile Slc7a7 knockout mice demonstrated 70% lower body weights, 87% lower plasma IGF-1 concentrations and delayed skeletal development. Because poor survival prevents evaluation of mature knockout mice, we generated a conditional Slc7a7 deletion in mature osteoblasts or mesenchymal cells of the osteo-chondroprogenitor lineage, but no differences in bone architecture were observed. Overall, global Slc7a7 deficiency caused growth failure with low plasma IGF-1 concentrations and delayed skeletal development, but Slc7a7 deficiency in the osteoblastic lineage was not a major contributor to these phenotypes. Future studies utilizing additional tissue-specific Slc7a7 knockout models may help dissect cell-autonomous and non-cell-autonomous mechanisms underlying phenotypes in LPI.

Healthcare utilisation in the last year of life in internal medicine, young-old versus old-old.

BACKGROUND: With increasing cost of healthcare in our aging society, a consistent pain point is that of end-of-life care. It is particularly difficult to prognosticate in non-cancer patients, leading to more healthcare utilisation without improving quality of life. Additionally, older adults do not age homogenously. Hence, we seek to characterise healthcare utilisation in young-old and old-old at the end-of-life. METHODS: We conducted a single-site retrospective review of decedents under department of Advanced Internal Medicine (AIM) over a year. Young-old is defined as 65-79 years; old-old as 80 years and above. Data collected was demographic characteristics; clinical data including Charlson Comorbidity Index (CCI), FRAIL-NH and advance care planning (ACP); healthcare utilisation including days spent in hospital, hospital admissions, length of stay of terminal admission and clinic visits; and quality of end-of-life care including investigations and symptomatic control. Documentation was individually reviewed for quality of communication. RESULTS: One hundred eighty-nine older adult decedents. Old-old decedents were mostly females (63% vs. 42%, p = 0.004), higher CCI scores (7.7 vs 6.6, p = 0.007), similarly frail with lower polypharmacy (62.9% vs 71.9%, p = 0.01). ACP uptake was low in both, old-old 15.9% vs. young-old 17.5%. Poor prognosis was conveyed to family, though conversation did not result in moderating extent of care. Old-old had less healthcare utilisation. Adjusting for sex, multimorbidity and frailty, old-old decedents had 7.3 ± 3.5 less hospital days in their final year. Further adjusting for cognition and residence, old-old had 0.5 ± 0.3 less hospital admissions. When accounted for home care services, old-old spent 2.7 ± 0.8 less hospital days in their last admission. CONCLUSION: There was high healthcare utilisation in older adults, but especially young-old. Enhanced education and goal-setting are needed in the acute care setting. ACP needs to be reinforced in acute care with further research to evaluate if it reduces unnecessary utilisation at end-of-life.

WITHDRAWN: Can Traditional Chinese Medicine provide insights into controlling the COVID-19 pandemic: Serpentinization-induced lithospheric long-wavelength magnetic anomalies in Proterozoic bedrocks in a weakened geomagnetic field mediate the aberrant transformation of biogenic molecules in COVID-19 via magnetic catalysis.

This article has been withdrawn at the request of the authors and the editors. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/our-business/policies/article-withdrawal.

Development of humanized mouse and rat models with full-thickness human skin and autologous immune cells.

The human skin is a significant barrier for protection against pathogen transmission. Rodent models used to investigate human-specific pathogens that target the skin are generated by introducing human skin grafts to immunocompromised rodent strains. Infection-induced immunopathogenesis has been separately studied in humanized rodent models developed with human lymphoid tissue and hematopoietic stem cell transplants. Successful co-engraftment of human skin, autologous lymphoid tissues, and autologous immune cells in a rodent model has not yet been achieved, though it could provide a means of studying the human immune response to infection in the human skin. Here, we introduce the human Skin and Immune System (hSIS)-humanized NOD-scid IL2Rγnull (NSG) mouse and Sprague-Dawley-Rag2tm2hera Il2rγtm1hera (SRG) rat models, co-engrafted with human full-thickness fetal skin, autologous fetal lymphoid tissues, and autologous fetal liver-derived hematopoietic stem cells. hSIS-humanized rodents demonstrate the development of human full-thickness skin, along with autologous lymphoid tissues, and autologous immune cells. These models also support human skin infection following intradermal inoculation with community-associated methicillin-resistant Staphylococcus aureus. The co-engraftment of these human skin and immune system components into a single humanized rodent model could provide a platform for studying human skin infections.

Moving beyond the mousetrap: current and emerging humanized mouse and rat models for investigating prevention and cure strategies against HIV infection and associated pathologies.

The development of safe and effective combination antiretroviral therapies for human immunodeficiency virus (HIV) infection over the past several decades has significantly reduced HIV-associated morbidity and mortality. Additionally, antiretroviral drugs have provided an effective means of protection against HIV transmission. Despite these advances, significant limitations exist; namely, the inability to eliminate HIV reservoirs, the inability to reverse lymphoid tissues damage, and the lack of an effective vaccine for preventing HIV transmission. Evaluation of the safety and efficacy of therapeutics and vaccines for eliminating HIV reservoirs and preventing HIV transmission requires robust in vivo models. Since HIV is a human-specific pathogen, that targets hematopoietic lineage cells and lymphoid tissues, in vivo animal models for HIV-host interactions require incorporation of human hematopoietic lineage cells and lymphoid tissues. In this review, we will discuss the construction of mouse models with human lymphoid tissues and/or hematopoietic lineage cells, termed, human immune system (HIS)-humanized mice. These HIS-humanized mouse models can support the development of functional human innate and adaptive immune cells, along with primary (thymus) and secondary (spleen) lymphoid tissues. We will discuss applications of HIS-humanized mouse models in evaluating the safety and efficacy of therapeutics against HIV reservoirs and associated immunopathology, and delineate the human immune response elicited by candidate HIV vaccines. In addition to focusing on how these HIS-humanized mouse models have already furthered our understanding of HIV and contributed to HIV therapeutics development, we discuss how emerging HIS-humanized rat models could address the limitations of HIS-mouse models.

Prevalence of Low Back Pain, Pelvic Girdle Pain, and Combination Pain in a Postpartum Ontario Population.

OBJECTIVE: The purpose of this study was to determine the point prevalence of low back pain (LBP), pelvic girdle pain (PGP), and/or combination pain (COMBO pain) and period prevalence (presence or absence of any of those pains), as well as to identify risk factors at 1, 3, and 6 months postpartum in a Canadian population. METHODS: Participants from a previous pregnancy study participated in a postpartum survey administered over the telephone at 1, 3, and 6 months following delivery. The survey included questions about LBP, PGP, or COMBO pain during the postpartum period, as well as questions related to risk factors (Canadian Task Force Classification II-3). RESULTS: At 1, 3, and 6 months postpartum, responses from 46, 58, and 64 participants, respectively, demonstrated that 15%-21% of women experienced LBP and up to 4% of women experienced COMBO pain (point prevalence). At no time point was PGP reported to occur alone. Period prevalence of back pain for the participants returned to pre-pregnancy levels at each time point. Back pain during pregnancy was the only risk factor identified for back pain at 3 and 6 months postpartum. CONCLUSION: This study demonstrated that 76% to 80% of respondents were pain free at 1, 3, and 6 months postpartum. Pregnancy-related back pain was the only risk factor associated with postpartum-related pain at the 1 to 3 and 3 to 6 month time interval. Identification of site-specific postpartum-related back pain may assist in determination of management and treatment plans for this population.

Conditional Spike Transmission Mediated by Electrical Coupling Ensures Millisecond Precision-Correlated Activity among Interneurons In Vivo.

Many GABAergic interneurons are electrically coupled and in vitro can display correlated activity with millisecond precision. However, the mechanisms underlying correlated activity between interneurons in vivo are unknown. Using dual patch-clamp recordings in vivo, we reveal that in the presence of spontaneous background synaptic activity, electrically coupled cerebellar Golgi cells exhibit robust millisecond precision-correlated activity which is enhanced by sensory stimulation. This precisely correlated activity results from the cooperative action of two mechanisms. First, electrical coupling ensures slow subthreshold membrane potential correlations by equalizing membrane potential fluctuations, such that coupled neurons tend to approach action potential threshold together. Second, fast spike-triggered spikelets transmitted through gap junctions conditionally trigger postjunctional spikes, depending on both neurons being close to threshold. Electrical coupling therefore controls the temporal precision and degree of both spontaneous and sensory-evoked correlated activity between interneurons, by the cooperative effects of shared synaptic depolarization and spikelet transmission.

Implementation of directly observed patient hand hygiene for hospitalized patients by hand hygiene ambassadors in Hong Kong.

BACKGROUND: The importance of compliance with hand hygiene by patients is increasingly recognized to prevent health care-associated infections. METHODS: This descriptive study observed the effects of an education campaign, targeted to increase patients' self-initiated hand hygiene, and a hand hygiene ambassador-initiated directly observed hand hygiene program on patients' hand hygiene compliance in a university-affiliated hospital. RESULTS: The overall audited compliance of patients' self-initiated hand hygiene was only 37.5%, with a rate of 26.9% (112/416 episodes) before meals and medications, 27.5% (19/69 episodes) after using a urinal or bedpan, and 89.7% (87/97 episodes) after attending toilet facilities. Patients referred from a residential care home for older adults had significantly lower hand hygiene compliance (P = .007). Comparatively, the overall audited compliance of ambassador-initiated directly observed hand hygiene was 97.3% (428/440 episodes), which was significantly higher than patients' self-initiated hand hygiene via a patient education program (37.5%, 218/582 episodes, P < .001). CONCLUSIONS: Directly observed hand hygiene can play an important role in improving compliance with hand hygiene by hospitalized patients.

Spontaneous Coronary Dissection Masquerading as Benign Fascicular Ventricular Tachycardia.

Spontaneous coronary artery dissection is an uncommon cause of acute coronary syndrome. Diagnosis of coronary artery dissection is made on coronary angiogram and prompt revascularisation is the key in management. We present a case of coronary artery dissection with an atypical presentation of cardiac arrhythmia mimicking benign fascicular ventricular tachycardia. A high index of suspicion and early coronary angiogram allowed us to diagnose and treat this potentially life-threatening disease.

Effect of proactive infection control measures on benchmarked rate of hospital outbreaks: An analysis of public hospitals in Hong Kong over 5 years.

BACKGROUND: Hospital outbreaks of epidemiologically important pathogens are usually caused by lapses in infection control measures and result in increased morbidity, mortality, and cost. However, there is no benchmark to compare the occurrence of hospital outbreaks across hospitals. METHODS: We implemented proactive infection control measures with an emphasis on timely education of health care workers and hospitalized patients at Queen Mary Hospital, a teaching hospital. Our benchmarked performance (outbreak episodes per 1 million patient discharges and 1 million patient-days) was compared with those of other regional public hospitals without these additional proactive measures in place between 2010 and 2014. RESULTS: During the study period, Queen Mary Hospital had 1 hospital outbreak resulting in 1.48 and 0.45 outbreak episodes per 1 million patient discharges and patient-days, respectively, values significantly lower than the corresponding overall rates in the 7 acute regional hospitals (24.26 and 6.70 outbreak episodes per 1 million patient discharges and patient-days, respectively; P < .001) and that of all 42 public hospitals in Hong Kong (41.62 and 8.65 outbreak episodes per 1 million patient discharges and patient-days, respectively; P < .001). CONCLUSIONS: The results of this large study on benchmarked rate of hospital outbreaks per patient discharges or patient-days suggests that proactive infection control interventions may minimize the risk of hospital outbreaks.

Upstream distal regulatory elements contact the Lmo2 promoter in mouse erythroid cells.

The Lim domain only 2 (Lmo2) gene encodes a transcriptional cofactor critical for the development of hematopoietic stem cells. Several distal regulatory elements have been identified upstream of the Lmo2 gene in the human and mouse genomes that are capable of enhancing reporter gene expression in erythroid cells and may be responsible for the high level transcription of Lmo2 in the erythroid lineage. In this study we investigate how these elements regulate transcription of Lmo2 and whether or not they function cooperatively in the endogenous context. Chromosome conformation capture (3C) experiments show that chromatin-chromatin interactions exist between upstream regulatory elements and the Lmo2 promoter in erythroid cells but that these interactions are absent from kidney where Lmo2 is transcribed at twelve fold lower levels. Specifically, long range chromatin-chromatin interactions occur between the Lmo2 proximal promoter and two broad regions, 3-31 and 66-105 kb upstream of Lmo2, which we term the proximal and distal control regions for Lmo2 (pCR and dCR respectively). Each of these regions is bound by several transcription factors suggesting that multiple regulatory elements cooperate in regulating high level transcription of Lmo2 in erythroid cells. Binding of CTCF and cohesin which support chromatin loops at other loci were also found within the dCR and at the Lmo2 proximal promoter. Intergenic transcription occurs throughout the dCR in erythroid cells but not in kidney suggesting a role for these intergenic transcripts in regulating Lmo2, similar to the broad domain of intergenic transcription observed at the human ß-globin locus control region. Our data supports a model in which the dCR functions through a chromatin looping mechanism to contact and enhance Lmo2 transcription specifically in erythroid cells. Furthermore, these chromatin loops are supported by the cohesin complex recruited to both CTCF and transcription factor bound regions.

Introduction of an electronic monitoring system for monitoring compliance with Moments 1 and 4 of the WHO "My 5 Moments for Hand Hygiene" methodology.

BACKGROUND: MedSense is an electronic hand hygiene compliance monitoring system that provides Infection Control Practitioners with continuous access to hand hygiene compliance information by monitoring Moments 1 and 4 of the WHO "My 5 Moments for Hand Hygiene" guidelines. Unlike previous electronic monitoring systems, MedSense operates in open cubicles with multiple beds and does not disrupt existing workflows. METHODS: This study was conducted in a 6-bed neurosurgical intensive care unit with technical development and evaluation phases. Healthcare workers (HCWs) wore an electronic device in the style of an identity badge to detect hand hygiene opportunities and compliance. We compared the compliance determined by the system and an infection control nurse. At the same time, the system assessed compliance by time of day, day of week, work shift, professional category of HCWs, and individual subject, while the workload of HCWs was monitored by measuring the amount of time they spent in patient zones. RESULTS: During the three-month evaluation phase, the system identified 13,694 hand hygiene opportunities from 17 nurses, 3 physiotherapists, and 1 healthcare assistant, resulting in an overall compliance of 35.1% for the unit. The per-indication compliance for Moment 1, 4, and simultaneous 1 and 4 were 21.3% (95%CI: 19.0, 23.6), 39.6% (95%CI: 37.3, 41.9), and 49.2% (95%CI: 46.6, 51.8), respectively, and were all statistically significantly different (p < 0.001). In the four 20-minute sessions when hand hygiene was monitored concurrently by the system and infection control nurse, the compliance were 88.9% and 95.6% respectively (p = 0.34), and the activity indices were 11.1 and 12.9 opportunities per hour, respectively. The hours from 12:00 to 14:00 had a notably lower compliance (21.3%, 95%CI: 17.2, 25.3) than nearly three quarters of the other periods of the day (p < 0.001). Nurses who used shared badges had significantly (p < 0.01) lower compliance (23.7%, 95%CI: 17.8, 29.6) than both the registered nurses (36.1%, 95%CI: 34.2, 37.9) and nursing officers (34.0%, 95%CI: 31.1, 36.9) who used named badges. CONCLUSION: MedSense provides an unobtrusive and objective measurement of hand hygiene compliance. The information is important for staff training by the infection control team and allocation of manpower by hospital administration.

Promotion of seasonal influenza vaccination among staff in residential care homes for elderly in Hong Kong.

Annual influenza epidemics continue to cause worldwide morbidity, mortality and societal disruption, especially among the aged residents of residential care homes for the elderly (RCHEs). Vaccination remains the most effective measure to prevent influenza and its associated complications. The seasonal influenza vaccine uptake rates among RCHE staff were much lower than that among residents. In order to increase uptake of influenza vaccination among RCHE staff in Hong Kong, this study developed and evaluated amultimodal vaccine promotion program (VPP) based on identified factors affecting vaccination acceptance or refusal within the Hong Kong Chinese context. Vaccine acceptance was found to be significantly associated with belief in vaccine efficacy, duration of service, staff group and providing direct care to residents. The focus group study revealed that RCHE staff's belief in the efficacy and safety of the vaccine played a major role in vaccine acceptance. VPP effectiveness was evaluated with a cluster randomised controlled trial among RCHEs with staff vaccination rates below 50%. Compared with 2008/09, the 2009/10 mean staff vaccination rates increased significantly in both the intervention (39.4% to 59.6% (P < 0.001)) and control groups (36.3% to 47.6% (P = 0.008)). RCHE staff in the intervention group had a higher vaccination rate than in the control group (59.6% versus 47.6%, P = 0.072). This program reinforces the importance of a comprehensive and culturally sensitive approach to promote influenza vaccination for RCHE staff.

Encoding of oscillations by axonal bursts in inferior olive neurons.

Inferior olive neurons regulate plasticity and timing in the cerebellar cortex via the climbing fiber pathway, but direct characterization of the output of this nucleus has remained elusive. We show that single somatic action potentials in olivary neurons are translated into a burst of axonal spikes. The number of spikes in the burst depends on the phase of subthreshold oscillations and, therefore, encodes the state of the olivary network. These bursts can be successfully transmitted to the cerebellar cortex in vivo, having a significant impact on Purkinje cells. They enhance dendritic spikes, modulate the complex spike pattern, and promote short-term and long-term plasticity at parallel fiber synapses in a manner dependent on the number of spikes in the burst. Our results challenge the view that the climbing fiber conveys an all-or-none signal to the cerebellar cortex and help to link learning and timing theories of olivocerebellar function.