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CARM1 is an arginine methyltransferase that has crucial roles in a number of cellular pathways and is being explored as a therapeutic target in diseases such as cancer and neurodegenerative disorders. Its deregulation at the protein level was found to have potential prognostic value, and as such, its protein levels are regularly assessed through the common practice of western blotting (WB). Our group uncovered that CARM1 has biochemical properties that complicate its analysis by standard WB sample preparation techniques. Here, we show that CARM1 has the ability to form SDS-resistant aggregates that effectively hinder gel migration in SDS-PAGE. CARM1 levels and the temperature at the denaturation step can both influence CARM1 aggregation, which prompts the use of additional measures to ensure representative detection at the protein level. We have demonstrated the formation of CARM1 aggregates in both cell and tissue extracts, making these findings an important consideration for any CARM1-related study. We also show how aggregate formation in models of CARM1 overexpression can hinder proteomic studies. Having identified factors that can induce CARM1 aggregation, we suggest alternative sample preparation techniques that allow for clear resolution of the protein in stringent denaturing conditions while avoiding aggregation., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

The Health App Review Tool (HART) is a novel assessment designed to match users with Alzheimer's disease or related dementias (ADRD) and caregivers to mobile applications that support health and wellness. The objectives of this study were to gather stakeholder feedback on the HART and then to implement revisions. Thirteen participants completed in-depth Think Aloud interviews. Participants shared qualitative feedback on each HART item. Participant feedback was analyzed via in-depth video-audio review. Feedback was implemented as actionable HART revisions. On average, the majority of participants rated items as "adequate"; however, qualitative findings indicated the need for improvement in conciseness, clarity, and understandability. Conciseness was addressed by combining related concepts into multi-items, clarity through the addition of specific examples, and understandability through improved verbiage. The HART has been refined from 106 items to 17 items through extensive revisions to the clarity, conciseness, and explanations provided throughout the assessment.

Purpose: Despite the benefits of wheelchair-mounted robotic arms (WMRAs), occupational therapists are not yet widely involved in the recommendation or implementation of these assistive devices. The purpose of this study was to investigate and compare the current practices and perspectives of occupational therapists who had and had not recommended a WMRA on the recommendation, training, and implementation of WMRAs., Methods: This was a descriptive cross-sectional study. An online survey was sent to Canadian, European, and American occupational therapists who had or had not worked with WMRAs. Respondents were asked close-ended questions about their experience, role, barriers, motivations, and future needs regarding WMRAs. We compared results between respondents who had and had not recommended WMRAs using descriptive statistics., Results: Ninety-three North American and European occupational therapists completed the survey. Of those, 29 (31.2%) had recommended a WMRA, mostly the JACO robotic arm ( n  = 26, 89.7%) in rehabilitation centres ( n  = 18, 62.1%). Their perspectives on their role and barriers related to WMRAs were similar to those who had never recommended a WMRA. All respondents recognised the relevance of occupational therapists' contribution, and most reported interest in WMRAs ( n  = 76, 81.7%). However, many barriers emerged, mainly related to limited funding ( n  = 49, 76.6%), lack of training and knowledge ( n  = 38, 59.4%), and resource constraints ( n  = 37, 54.4%). Future needs identified matched these barriers., Conclusion: This survey provides novel insight into occupational therapists' perspectives on WMRAs. It highlights that health professionals need to have easier access to funding, formal training, and resources to support their involvement with WMRAs.Implications for rehabilitationMost occupational therapists are interested in working with WMRAs, considering the potential of these devices to support individuals with upper extremity impairments in their daily activities. They also recognise their unique contribution to the assessment, recommendation, and implementation process among multidisciplinary teams.WMRA recommendation is relevant in various clinical settings and with a wide range of client populations. Nevertheless, it appears that occupational therapists working with adults, in rehabilitation centres or specialised clinics, may have more opportunities to get involved in this process and to attend formal training on this technology, as compared to other settings.Many barriers remain, impeding occupational therapists' role in the recommendation and implementation of WMRAs. Addressing these barriers may increase the number of devices that are successfully adopted and utilised by individuals with upper extremity impairments. In particular, future research and health policies should focus on access to sufficient funding, formal training, and resources for occupational therapists relative to their role in recommending and implementing WMRAs.

Purpose: Although approximately 45% of adults with Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS) are permanent wheelchair users, this sub population has been less studied. The purpose of this study was to document wheelchair mobility, motor performance, and participation in a cohort of adult wheelchair users with ARSACS., Methods: We recruited 36 manual and powered wheelchair users with ARSACS, aged between 34 and 64 years, for this cross-sectional study. Participants completed measures regarding wheelchair mobility (Wheelchair Skills Test Questionnaire [WST-Q-F], Wheelchair Use Confidence Scale [WheelCon-F] and Wheelchair Outcome Measure [WhOM-F]), motor performance (Scale for the Assessment and Rating of Ataxia [SARA], Disease Severity Index for adults with ARSACS [DSI-ARSACS], Upper Extremity Performance Test for the Elderly [TEMPA], Standardised Finger to Nose Test [SFNT], grip strength, pinch strength, Lower Extremity Motor Coordination Test [LEMOCOT], Berg Balance Scale [BBS], Timed Up and Go [TUG] and 10-meter Walk Test [10mWT]), and participation (Barthel Index, LSA-F and LIFE-H). Results were compared between age groups (≤49 years and ≥50 years), types of wheelchair used, and available reference values. Correlations were computed between wheelchair mobility, upper limb function, and participation., Results: Participants presented limitations regarding wheelchair skills, motor performance, and participation in daily activities. Despite preserved upper limb strength, wheelchair skills, upper and lower limb coordination, standing balance, and functional independence were generally more impaired after 50 years of age and among powered wheelchair users. Significant moderate correlations were found between wheelchair skills and self-efficacy, upper limb strength and coordination, and participation in daily and social activities., Conclusions: This study provided the first data sets describing specific characteristics of manual and powered wheelchair users with ARSACS. It supports a need to offer wheelchair skills training interventions to adults with ARSACS, which could increase their daily and social participation.IMPLICATIONS FOR REHABILITATIONAdult wheelchair users with ARSACS present with limited wheelchair skills, significantly impaired motor performance, and reduced participation that generally decreases with age. This profile may serve as comparative data for clinicians to anticipate disease progression.This study provides the first data on distinguishing characteristics between PWC users and MWC users with ARSACS. The main characteristics of PWC users include more severe functional limitations and motor impairments, as well as limited grip strength that contrasts with the general preservation of this function among other adults with ARSACS.There is a need to offer and evaluate wheelchair skills training interventions in the future for adults with ARSACS. The general preservation of grip and pinch strength observed in this population suggests a potential for improvement. Considering the associations found between wheelchair mobility and participation, such interventions may increase users' daily and social participation.

Purpose: To establish the reliability and construct validity of two French-Canadian versions of assessment tools for manual (MWC) and powered wheelchair (PWC) users with autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS): the Wheelchair Skills Test Questionnaire (WST-Q-F) version 5.0 and the Wheelchair Use Confidence Scale (WheelCon-F) Short Form., Methods: We recruited 32 MWC and PWC users with ARSACS aged between 34 and 64 years. Participants completed measures twice within 2 weeks for test-retest reliability and to determine the standard error of measurement. Construct validity was established by verifying hypothesized relationships between wheelchair use scores and other variables regarding personal factors, body functions, and activities. Participants' scores were also compared with those of MWC and PWC users with other diagnoses to explore known-groups validity., Results: Adequate to excellent test-retest reliability values were found for all questionnaires (intraclass correlation coefficients between 0.506 and 0.995). SEMs were acceptable, ranging from 3.3 to 9.1 on the WST-Q-F and from 0.7 to 1.2 on the WheelCon-F. Moderate to excellent correlations supported construct validity for the WST-Q-F and the WheelCon-F. Compared with other populations, adults with ARSACS reported limited wheelchair skills, but similar wheelchair confidence., Conclusions: The WST-Q-F and the WheelCon-F have excellent test-retest reliability, acceptable measurement errors, and support for construct validity in adults with ARSACS. This study was the first to evaluate these assessment tools in this population, and the results support a need to implement wheelchair skills training interventions in this population.IMPLICATIONS FOR REHABILITATIONThe WST-Q-F version 5.0 and the WheelCon-F Short Form are reliable and valid assessment tools to measure manual and powered wheelchair skills and self-efficacy in adults with autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS).The WST-Q-F version 5.0 and the WheelCon-F Short Form can be used to evaluate and describe wheelchair skills and wheelchair use self-efficacy in clinical practice and are recommended for future research in adults with ARSACS. These outcome tools may be especially useful to measure the effects of a wheelchair training program.

Background: States of chronic overnutrition and undernutrition are both associated with impaired bone health and increased fracture risk but there are no data on bone microarchitecture following short-term controlled nutritional challenges., Objective: The purpose of our study was to evaluate the impact of short-term high-caloric feeding and fasting on bone microarchitecture. We hypothesized that both high-caloric feeding and fasting would have negative effects on microarchitecture., Materials and Methods: We recruited 23 adult healthy subjects (13 males, 10 females, mean age 33.2 ± 1.4 years, mean BMI 26.0 ± 1.5 kg/m 2 ). Subjects underwent an in-patient 10-day high-caloric visit (caloric intake with goal to achieve 7% weight gain), after which they went home to resume a normal diet for 13-18 days (stabilization period), and were then readmitted for a 10-day in-patient fasting stay (no caloric intake). All subjects underwent HRpQCT (XtremeCT, Scanco Medical AG, Brüttisellen, Switzerland) of the distal tibia and distal radius after each visit to assess volumetric bone mineral density (vBMD), trabecular and cortical microarchitecture, and strength estimates. The Wilcoxon signed rank test was used to perform within group comparisons., Results: During the high-caloric period, there was a mean increase in weight by 6.3 + 1.7% (p < 0.0001). There were no significant changes in bone parameters in the distal tibia or distal radius (p > 0.05). During the stabilization period there was a significant reduction in weight by -2.7 + 1.9% (p < 0.0001) but no change in bone parameters (p > 0.05). During the fasting period there was a further reduction in weight by -8.8 + 1.2% (p < 0.0001). In the distal tibia, there was a significant increase in total and cortical vBMD, trabecular and cortical parameters as well as strength estimates (p < 0.05). In the distal radius there was an increase in total and trabecular vBMD (p < 0.05), while there were no changes in other microarchitecture parameters or strengths estimates., Conclusion: Short-term fasting after high-caloric feeding improves vBMD, bone microarchitecture and strength estimates of the distal tibia, while short-term high-caloric feeding does not change vBMD or microarchitecture. These results suggest that short-term fasting after high-caloric feeding in healthy individuals improves bone health and that these changes can be detected using HRpQCT in-vivo., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The protein arginine methyltransferase family (PRMT) is known as being the catalytic driving force for arginine methylation. This specific type of post translational modification is extensively used in biological processes, and therefore is highly relevant in the pathology of a profusion of diseases. Since altered PRMT expression or deregulation has been shown to contribute to a vast range of those diseases including cancer, their study is of great interest. Although an increasing number of substrates are being discovered for each PRMT, large scale proteomic methods can be used to identify novel interactors/substrates, further elucidating the role that PRMTs perform in physiological or disease states. Here, we describe the use of affinity purification (AP) coupled with stable isotope labeling with amino acids in cell culture (SILAC) quantitative mass spectrometry (MS) to identify protein interactors and substrates of PRMTs. We also explore the possibility of exploiting the fact most PRMTs display lower dissociation rates with their hypomethylated substrates as a strategy to increase the proportion of substrates identified in AP/MS studies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019. Published by Elsevier Inc.)

Winter survival is a determinant factor for the persistence of perennials grown in northern climates. High winter survival cultivars, however, have lower yield due to their early transition into a dormant state in the fall. Here we describe a whole plant assay entirely performed indoor in growth chambers and walk-in freezers to identify low-dormant genotypes with superior freezing tolerance within populations of open pollinated species. Three successive freezing stresses are applied to a broad base of 3000 genotypes to progressively eliminate 97% of the population and to retain only the 3% best performing genotypes. This approach can be used to generate recurrently selected populations in different species.

Nanoparticles that readily penetrate mucosal layers are desirable for a variety of biomedical applications. Nevertheless, most nanoparticles tend to be immobilized in mucus via steric and/or adhesive interactions. Contrary to the established opinion that poly(vinyl alcohol) (PVA) is mucoadhesive, we discovered that coating otherwise mucoadhesive nanoparticles with certain partially hydrolyzed PVAs can aid particle mobility in mucus. We describe two approaches to producing such mucus-penetrating particles (non-covalent modification of pre-formed nanoparticles and emulsification in the presence of PVA) and provide mobility data in human cervicovaginal mucus ex vivo as measured by multiple particle tracking and bulk permeation. When coated with PVAs that are ≥95% hydrolyzed, nanoparticles as small as ~210nm were immobilized in mucus similarly to well-established mucoadhesive controls (P>0.05). However, nanoparticles coated with PVAs that are <95% hydrolyzed penetrated mucus with velocities significantly exceeding those for the mucoadhesive controls (P<0.001) and were mobile in the bulk permeation assay., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Most attempts to achieve sustained drug delivery to pulmonary tissues using nanoparticles have focused on mucoadhesive particles (MAP). However, MAP become trapped in the luminal mucus layer and, as a result, are largely eliminated from the respiratory tract by mucociliary escalator and expiratory clearance, which undermines their sustained release potential. Recent studies have shown that mucus-penetrating particles (MPP) engineered to diffuse through mucus can avoid rapid mucociliary clearance in vivo and persist in the lung longer. Nonetheless, it has not been confirmed that MPP encapsulating small molecules can sustain drug release in the lung longer than MAP of similar size and core composition. As a proof of concept, we encapsulated fluticasone propionate (FP) into poly(lactide)-based MPP and MAP (both ∼ 200 nm diameter, ∼ 30-35% drug loading) and evaluated their pulmonary residence by measuring FP levels in mouse lungs over 24h following intratracheal instillation. Furthermore, we evaluated the duration of action of FP MPP in a rat lung inflammation model compared to that of a non-encapsulated FP control. In rodents, pulmonary delivery of FP formulated as MPP provided a 60% higher local exposure compared to MAP and extended the single dose efficacy by at least 16 h compared to non-encapsulated FP., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Purpose: Traditional polymeric nanoparticle formulations for prolonged local action during inhalation therapy are highly susceptible to muco-ciliary clearance. In addition, polymeric carriers are typically administered in high doses due to finite drug loading. For toxicological reasons, these carriers and their degradation byproducts are undesirable for inhalation therapy, particularly for chronic use, due to potential lung accumulation., Methods: We synthesized a novel, insoluble prodrug (MRPD) of a time-dependent β-lactam, meropenem, and formulated MRPD into mucus-penetrating crystals (MRPD-MPCs). After characterizing their mucus mobility (in vitro) and stability, we evaluated the lung pharmacokinetics of intratracheally-instilled MRPD-MPCs and a meropenem solution in guinea pigs., Results: Meropenem levels rapidly declined in the lungs of guinea pigs receiving meropenem solution compared to those given MRPD-MPCs. At 9 h after dosing, drug levels in the lungs of animals that received meropenem solution dropped to 12 ng/mL, whereas those that received MRPD-MPCs maintained an average drug level of ≥1,065 ng/mL over a 12-h period., Conclusions: This work demonstrated that the combination of prodrug chemistry and mucus-penetrating platform created nanoparticles that produced sustained levels of meropenem in guinea pig lungs. This strategy represents a novel approach for sustained local drug delivery to the lung without using encapsulating matrices.

We show how to realize fast and high-fidelity quantum nondemolition qubit readout using longitudinal qubit-oscillator interaction. This is accomplished by modulating the longitudinal coupling at the cavity frequency. The qubit-oscillator interaction then acts as a qubit-state dependent drive on the cavity, a situation that is fundamentally different from the standard dispersive case. Single-mode squeezing can be exploited to exponentially increase the signal-to-noise ratio of this readout protocol. We present an implementation of this longitudinal parametric readout in circuit quantum electrodynamics and a possible multiqubit architecture.

Introduction: Topical ophthalmic formulations of corticosteroids are commonly used to treat a variety of ocular diseases and conditions that have an inflammatory component. The purpose of this study was to evaluate the effect of the mucus-penetrating particle (MPP) technology on the pharmacokinetic profile of loteprednol etabonate in the ocular tissues of rabbits., Methods: Forty-eight New Zealand White rabbits were randomly assigned to two groups (n = 3 rabbits or 6 eyes per time point) and treated with either the novel loteprednol etabonate MPP suspension formulation, 0.4% (LE-MPP 0.4%), or the commercial Lotemax(®)-brand loteprednol etabonate ophthalmic suspension, 0.5% (Lotemax 0.5%) (Bausch & Lomb Incorporated, Inc., Rochester, NY, USA). Samples of aqueous humor, various ocular tissues, and plasma were collected from animals over a 12-h period after a single dose of the test articles. Loteprednol etabonate concentrations were assayed using liquid chromatography-tandem mass spectrometry (LC/MS/MS)., Results: Loteprednol etabonate was rapidly absorbed into ocular tissues following administration of either formulation. A higher ocular exposure was achieved using LE-MPP 0.4%, with peak concentrations of approximately threefold higher in ocular tissues and the aqueous humor than Lotemax 0.5%., Conclusions: Administration of LE-MPP 0.4% improved loteprednol etabonate pharmacokinetic profile in ocular tissues of rabbits. The results of this study support the premise that the MPP technology can be used to enhance ocular exposure for topically applied therapeutic agents. Further studies to assess the clinical efficacy and safety of the LE-MPP formulation are warranted.

Freezing tolerance is a determinant factor of persistence of perennials grown in northern climate. Selection for winterhardiness in field nurseries is difficult because of the unpredictability of occurrence of test winters allowing the identification of hardy genotypes. Here we describe a whole-plant assay entirely performed indoor in growth chambers and walk-in freezers to identify genotypes with superior tolerance to freezing within populations of open pollinated species. Three successive freezing stresses are applied to progressively eliminate 90 % of the population and to retain only the 10 % best performing genotypes. This approach can be used to generate recurrently selected populations more tolerant to freezing in different species.

We show that controlling relative phases of electromagnetic fields driving an atom with a Δ-configuration energy-level structure enables optical susceptibility to be engineered in novel ways. In particular, relative-phase control can yield electromagnetically induced transparency but with the benefit that the transparency window is sandwiched between an absorption and an amplification band rather than between two absorption bands in typical electromagnetically induced transparency. We show that this new phenomenon is achievable for a microwave field interacting with a fluxonium superconducting circuit.

Integrity profiling of HTS hits is valuable for verification of the hit identity and purity. This provides early discovery researchers with more confident SAR theories. Methodology for integrity profiling of HTS hits must be high throughput, consume little material, and selectively provide structure-based data. Analytical techniques that can be utilized for integrity profiling methods are reviewed for their appropriateness in sample preparation, component separation, detection, purity quantitation, identity confirmation, and follow-up.

This study was undertaken to confirm the presence of CCK receptor subtypes in calf pancreas and establish their cellular localization. Using specific antibodies against CCKA and CCKB receptors, somatostatin, glucagon and insulin, we were able to confirm by Western blot the presence of both CCK receptor protein subtypes in the calf pancreas as a 80-85-kDa CCKA receptor and 40-45-kDa CCKB receptor. By immunofluorescence, the CCKB receptor colocalizes with the islets' somatostatin delta cells, confirming what was previously shown in other species, as well as on ductal cells. We could not reproduce in the calf its colocalization with glucagon alpha cells as observed in human and rat. Any specific localization of CCKA receptors with our multiple antibodies failed. Our observation that the CCKB receptor subtype is specifically localized on pancreatic delta cells as well as on ductal cells lets us support the hypothesis that in this species, CCK could be involved in somatostatin metabolism as well as hydrelatic secretion; its effect on enzyme secretion would be indirect., (Copyright 2002 Elsevier Science B.V.)

Positive signs of pancreas regeneration were observed in rats after induced pancreatitis and partial pancreatectomy (1,2). Although the human pancreas did not regenerate after partial anatomic resection (3), the pig pancreas exhibited growth responses to bombesin after partial pancreatectomy (4). This study was undertaken to establish the time course of pancreatic inflammation, apoptosis, and hypertrophy and/or hyperplasia after partial pancreatectomy in pigs.

In rodents, cholecystokinin (CCK) induces pancreatic enzyme secretion and pancreas growth through its CCK(A) receptors. It is unknown whether occupation of the CCK(B) receptors present in pig and human pancreas can cause the same effects. This study evaluates CCK(B) receptor expression in rat, mouse, pig, and fetal human pancreata using Northern blot, Western blot, and immunofluorescence techniques. The reported 2.7-kb CCK(B) receptor mRNA transcript in the rat brain and gastric fundus is absent in pancreas; the message was, however, detected by RT-PCR and by a CCK(B) receptor antibody as an 80-kDa protein present uniquely in islet delta-cells. Proteins of 50 and 80 kDa appear in mouse pancreas, and proteins of 50 and 115 kDa appear in pig and human pancreas, respectively, all localized in islet delta-cells. Gastrin mRNAs are strongly present in fetal rat pancreas, and the hormone is localized in islets; both are repressed 10 days after birth. In conclusion, the CCK(B) receptors are present in pancreas of four species with exclusive location in islet delta-cells. In such a location, they could be indirectly involved in the control of enzyme secretion.

This study was undertaken to characterize the biochemical properties of rat pancreatic phospholipase D (PLD). Based on Western blot analysis of pancreas subcellular fractions, PLD1 was detected as a protein of 120 kDa associated with the microsomal fraction, whereas PLD2 appeared as a 105-kDa protein enriched in the microvesicular fraction. In these fractions, a low level of PLD activity was measured with an exogenous substrate containing phosphatidylinositol-4,5-bisphosphate (PIP2), unresponsive to guanosine triphosphate (GTP)gammaS and adenosine diphosphate (ADP)-ribosylation factor (ARF). Addition of unsaturated but not saturated fatty acids stimulated an oleate-dependent PLD activity that colocalized with the PLD1 enzyme in the crude plasma membrane and microsomal fractions. The transphosphatidylation reaction was maximal with either 200-400 mM (1.2-2.3%) ethanol or 25 mM (0.23%) 1-butanol, with an optimal pH between 6.5 and 7.2. Lipids extracted from the pancreatic membranes were potent inhibitors of the HL60 cell PLD activity when compared with those isolated from HL60 cells. Oleate-dependent PLD activity was less susceptible to these inhibitions. A phospholipase A1 (PLA1) activity hydrolyzing phosphatidylethanol also was found in the pancreatic membrane fractions and was nearly absent in the HL60 cells. This activity was completely inhibited by 400 nM tetrahydrolipstatin (THL), a lipase inhibitor. Pancreatic PLD1 and PLD2 activities could be measured after a chromatographic separation from microsomal membranes and high-speed supernatants, respectively. Activities of both enzymes were inhibited by oleate and required the presence of PIP2 in the substrate vesicles. ARF1 strongly activated PLD1 in a dose-dependent manner, and PLD2 was slightly responsive. Indirect immunofluorescence revealed that PLD2 is distributed throughout the pancreas, with a more intense staining in the islets. This study presents for the first time biochemical characteristics of the pancreatic PLD activities and shows the presence of oleate-dependent PLD1 and PLD2 activities, as well as PLD1 and PLD2 proteins in this gland.

Pancreatic growth occurs after CCK, CCK-induced pancreatitis, and pancreatectomy; the mechanisms involved remain unknown. This study evaluates mitogen-activated protein kinase (MAPK) activation and expression of cell cycle regulatory proteins after pancreatectomy to understand the cellular and molecular mechanisms involved in pancreas regeneration. Rats were killed 1-12 days after pancreatectomy, and p42/p44 MAPK activation, expression of the cyclins D and E, cyclin-dependent kinase (Cdk)-2 activity, retinoblastoma protein (pRb) hyperphosphorylation, and expression of the cyclin kinase inhibitors p15, p21, and p27 were examined. Pancreatic remnants exhibited sustained p42/p44 MAPK activation within 8 h. Cyclins D1 and E showed maximal expression after 2 and 6 days, coinciding with maximal hyperphosphorylation of pRb and Cdk2 activity. The expression of p15 vanished after 12 h, p27 disappeared gradually, and p21 increased early. The p27 complexed with Cdk2 dissociated after 2 days, whereas p21 associated in a reverse fashion. In conclusion, sustained activation of p42/p44 MAPKs and Cdk2 along with overexpression of cyclins D1 and E and reduction of p15 and p27 cyclin inhibitors occurred early after pancreatectomy and are active factors involved in signaling that leads to pancreas regeneration.

We have evaluated the presence and localization of the CCK(A) receptor in rat, mouse, pig and human fetal pancreas by Northern, Western blots and immunofluorescence techniques. In the rat, parallelism exists between development of the CCK(A) receptor mRNA and protein with maximal peaks of expression during the suckling period. In the course of pancreatitis induction, CCK(A) receptor mRNA were maximally expressed and sustained during the gland's regeneration. In the rat and mouse pancreas, the CCK(A) receptor protein is localized around the acinar cells and beta cells of the islets of Langerhans. In the adult pig and fetal human pancreas, the CCK(A) receptor proteins were detected by Western blot. By immunofluorescence, its detection was possible only in the islet of Langerhans of the pig pancreas. These new findings support the views that CCK plays important and various roles in specific physiological systems of the pancreas of different species., (Copyright 1999 Academic Press.)

A time-resolved optical (UV-vis) spectroscopic study of Roussin's black salt anion, Fe(4)S(3)(NO)(7)(-), revealed two separate intermediates, X and Y, following laser flash photolysis. Both intermediates react with nitric oxide with second-order kinetics to re-form the parent complex (k(NO)(X)() = 1.3 x 10(7) M(-)(1) s(-)(1); k(NO)(Y)() = 7.0 x 10(5) M(-)(1) s(-)(1) in aqueous solutions). The shorter-lived intermediate X was observed in time resolved infrared spectroscopic studies. Isotopic labeling experiments involving the exchange of black salt nitrosyls with (15)NO(2)(-) or (15)N(18)O were used to probe the correlation between nu(NO) bands and the anion structures. The identities of the intermediates are interpreted in terms of photolytic loss of chemically distinct nitrosyls found on the Fe(4)S(3)(NO)(7)(-) anion.

Described are studies directed at refining quantitative analysis of nitric oxide in solution using electrochemical techniques. The fabrication and behavior of several sensors based on modified carbon-based electrodes are reported. This technique has been used to resolve the vexing problem of determining the stoichiometry of the photochemical decomposition of the known antihypertension agent sodium nitroprusside, Na2[Fe(CN)5NO], as well as of two other metal nitrosyl complexes of biological interest, Roussin's black salt, NH4[Fe4S3(NO)7], and Roussin's red salt, Na2[Fe2S2(NO)4], in aqueous solutions. In this manner it was shown that the molar ratios of nitric oxide produced per starting complex photochemically decomposed were 0.95 +/- 0.03, 5.9 +/- 0.2, and 0.50 +/- 0.02 for Na2[Fe(C-N)5NO], NH4[Fe4S3(NO)7], and Na2[Fe2S2(NO)4], respectively.

Striatal and cortical projections arising from the central lateral thalamic nucleus were studied in rats by tracing the axons of small pools of neurons labeled anterogradely with biocytin. Cells of the central lateral nucleus have a morphology that conforms to the classic descriptions of the bushy cells which represent the main neuronal type of most thalamic nuclei. They display many short radiating dendrites studded with sessile spines, protrusions and grapelike appendages. The total extent of their dendritic fields is about 250 mu m. After leaving the nucleus, all central lateral axons course through the rostrolateral pole of the thalamic reticular nucleus, where they branch profusely, enter the striatum, where they distribute collaterals, and arborize in the motor cortex. At striatal level, central lateral fibers form a loosely organized network composed of varicose axonal branches that appear to contact en passant several striatal neurons. In the cortex. central lateral axons from multiple (four to five patches of terminations in layers Va and III aligned along the rostrocaudal extent of the motor area. The projection to layers I and II is very sparse, consisting of occasional branches which show few ramifications. Our results indicate that most, and perhaps all, central lateral relay neurons project to both the striatum and cerebral cortex. The patchy innervation of mid cortical layers of the frontal motor areas by central lateral afferents strongly argues against the nonspecific character of this projection. It is proposed that the central lateral nucleus, which receives a strong innervation from brainstem cholinergic afferents, takes part in a mechanism of attention related to the central initiation of directed patterns of movements.

Thalamostriatal projections arising from the posterior intralaminar nuclei (P1; the parafascicular nucleus and the adjacent caudalmost part of the posterior thalamic group) were studied in rats by tracing the axons of small pools of neurons labelled anterogradely with biocytin. Thirteen P1 cells were also stained by juxta cellular application of the tracer. Relay cells of P1 nuclei have a morphology that differs radically from the classical descriptions of the bushy cells which represent the main neuronal type of the sensory thalamic relay nuclei. P1 cells have ovoid or polygonal somata of approximately 20-25 microm, from which emerge four or five thick, long and poorly branched dendrites bearing spines and filamentous appendages; their dendritic domains extend for up to 1.5 mm. Before leaving the nucleus 20% of axons give off collaterals that ramify locally. All axons course through the thalamic reticular nucleus, where they also distribute collaterals, and arborize massively in the striatum and sparsely in the cerebral cortex. At the striatal level four or five collaterals leave the main axon and terminate in patches scattered dorsoventrally within a rostrocaudally oriented slab. As revealed by calbindin D-28k immunohistochemistry, only the matrix compartment receives terminations from P1 axons. The cortical branch form small terminal puffs centred upon layer VI of the motor cortex. Before entering the striatum some axons of the parafascicular nucleus give rise to descending collaterals that arborize in the entopeduncular nucleus, in the subthalamic nucleus and in the vicinity of the red nucleus. Other axons arising from the caudal part of the posterior group send descending branches only to the entopeduncular nucleus. These findings show that P1 cells belong to a distinct category of thalamic relay neurons which, beside their massive projection to the striatum, also distribute collaterals to other components of the basal ganglia. Moreover, these results provide the first direct evidence that virtually all P1 cells project to both striatum and cerebral cortex. Finally, it is proposed on the basis of morphological, histochemical and hodological criteria that the caudal part of the posterior thalamic group in the rat is homologous to the suprageniculate-limitans nuclei of cats and primates.

Thalamostriatal projections were studied in rats by tracing the axons of small pools of thalamic neurons labeled anterogradely with biocytin. Single-cell mapping of these projections revealed two types of thalamostriatal fibers. The first type arises from the bushy relay cells of the central lateral and associative thalamic nuclei which arborize sparsely in the striatum by means of long varicose axon collaterals. The second type of fiber arises from large, reticular-like, relay cells located in the parafascicular and ethmoid nuclei. These latter fibers form dense clusters of terminations within the striatum and they also send branches to other components of the basal ganglia. These different morphological features suggest that the two types of fibers subserve different functions.

This study investigates the pattern of axonal projections of single corticothalamic neurons from the rat primary visual cortex. Microiontophoretic injections of biocytin were made in cortical laminae V and VI to label small pools of corticothalamic cells and their intrathalamic axonal projections. After a survival period of 48 h, the animals were perfused and the tissue was processed for biocytin histochemistry. On the basis of the intrathalamic distribution of axonal fields and the types of terminations found in the thalamus, three types of corticothalamic projections were identified. (1) Cells of the upper part of lamina VI projected to the dorsal lateral geniculate nucleus where they arborized in rostrocaudally oriented bands or "rods" parallel to the lines of projection of retinal afferents. (2) Cells of the lower part of lamina VI projected to the lateral part of the lateral posterior nucleus and they also sent collaterals to the dorsal lateral geniculate nucleus where they participated in the formation of rods. (3) The corticothalamic projection of lamina V cells originated from collaterals of corticofugal cells whose main axons reached the tectum and/or the pontine nuclei. These collaterals never terminated within the dorsal lateral geniculate nucleus; they arborized in the lateral posterior, lateral dorsal and ventral lateral geniculate nuclei. All corticothalamic cells from lamina VI displayed the same type of axonal network made of long branches decorated by terminal boutons emitted "en passant" at the tip of fine stalks. Corticothalamic fibers arising from lamina V, however, generated varicose endings in restricted regions of their target nuclei. All corticothalamic axons derived from lamina VI cells, but not those derived from lamina V cells, gave off collaterals as they traversed the thalamic reticular complex. These results demonstrate that corticothalamic fibers arising from the rat primary visual cortex display a lamina-dependent projection pattern. In the light of previous studies on the topographical organization of corticothalamic projections, it is proposed that a similar organizational plan characterizes corticothalamic relationships in other sensory systems in the rat and in other species.

This study describes the axonal projections of single thalamic reticular (TR) neurons within the visual thalamus in rats. Experiments were performed under urethane anesthesia and reticular cells were labeled by extracellular or juxtacellular microiontophoretic applications of biocytin. The axonal arborizations of 19 TR cells projecting to the dorsal lateral geniculate nucleus (DLG) or to the lateral dorsal/lateral posterior complex (LD/LP) were reconstructed from serial horizontal sections. It was found that single TR cells projected within the limits of a single thalamic nucleus, either the DLG or the LD/LP complex, where their terminal fields formed rostrocaudally oriented rods (length: approximately 800 microns; diameter: approximately 100 microns) densely packed with grape-like boutons and varicosities. In addition, none of the labeled TR cells possessed recurrent axonal collaterals that ramified within the reticular complex itself. The functional implications of these morphological data for the synchronization of thalamic oscillations are discussed.

This study investigated the pattern of axonal projections of single corticothalamic neurons from the cortical barrel field representing the vibrissae in the rat. Microiontophoretic injections of biocytin were performed in cortical layers V and VI to label small pools of corticothalamic cells and their intrathalamic axonal projections. After a survival period of 48 h, the animals were perfused and the tissue was processed for biocytin histochemistry. On the basis of the intrathalamic distribution of axonal fields and of the types of terminations found in the thalamus, four types of corticothalamic projections were identified. (i) Cells of the upper part of layer VI projected exclusively to the ventral posteromedial (VPm) nucleus, where they arborized in long rostrocaudally oriented bands or 'rods'. (ii) All cells of the lower part of layer VI projected to the medial part of the thalamic posterior group (Pom) but the vast majority of them also collateralized in VPm where they participated in the formation of rods. (iii) A minority of corticothalamic cells in the lower portion of layer VI, possibly located under the interbarrel spaces (septae), arborized exclusively in Pom. (iv) The corticothalamic projection of layer V cells originated from collaterals of corticofugal cells whose main axons ran caudally towards the brainstem. These collaterals arborized exclusively in Pom or in the central lateral nucleus. All corticothalamic cells from layer VI displayed the same type of axonal network, made of long branches decorated by terminal buttons emitted en passant at the tip of fine stalks. Corticothalamic fibres arising from layer V pyramids, however, remained smooth as they ran across the lateral thalamus and they generated in Pom one or two clusters of large boutons. All corticothalamic axons derived from layer VI cells, but not those derived from layer V cells, gave off collaterals as they traversed the thalamic reticular complex. These observations are discussed in the light of previous studies bearing on the topological organization and function of corticothalamic projections to VPm and Pom in rats. The possibility that a similar cellular specificity and a similar organizational plan may characterize corticothalamic relationships in other sensory systems is also considered.

This study describes the axonal projections of single neurons of the thalamic reticular complex within the somatosensory thalamic nuclei in rats. Experiments were performed under urethane anaesthesia and reticular cells were labelled by extracellular microiontophoretic applications of biocytin. The axonal arborization of 25 thalamic reticular cells projecting to the ventrobasal (VB) nucleus and/or to the posterior thalamic (Po) complex were reconstructed from serial horizontal sections. Reticular cells labelled with biocytin display somatodendritic features similar to those reported previously. Their cell body is fusiform and their dendrites bear few spines and show a high degree of streaming along the horizontal curved axis of the nucleus. In most cells, axon-like beaded processes stem out from dendrites but, contrary to previous descriptions, no intrareticular axonal collateral was observed. The axonal arborization of most thalamic reticular cells is confined within the limits of a single thalamic nucleus; only two neurons were seen projecting to both the VB and the Po nuclei. In VB, termination fields form short rods (diameter approximately 150 microns, length approximately 200-300 microns) densely packed with grape-like boutons and varicosities; termination fields in Pro are larger, much less dense, and they are contained within a horizontal slab of tissue (thickness approximately 200 microns, mediolateral width approximately 400 microns, rostrocaudal length approximately 1 mm. By charting the position of all labelled cells within the thickness of the thalamic reticular complex, a strip-like arrangement was revealed. Cells projecting to Po occupy the innermost portion of the nucleus whereas those projecting to the ventral-posteromedial and ventral-posterolateral nuclei are located respectively in the middle and in the outer tiers of the nucleus. This strip-like reciprocity was confirmed by separate biocytin injections performed in VB and in Po. These results show that inhibition of reticular origin is distributed within the rat dorsal thalamus in a highly specific manner, most likely according to a principle of reciprocity within the somatotopic representation of the body.

The vast majority of corticothalamic (CT) axons projecting to sensory-specific thalamic nuclei arise from layer VI cells but intralaminar and associative thalamic nuclei also receive, to various degrees, a cortical input from layer V pyramidal cells. It is also well established that all long-range corticofugal projections reaching the brainstem and spinal cord arise exclusively from layer V neurons. These observations raise the possibility that the CT input from layer V cells may be collaterals of those long-range axons projecting below thalamic level. The thalamic projections of layer V cells were mapped at a single cell level following small microiontophoretic injections of biocytin performed in the motor, somatosensory and visual cortices in rats. Camera lucida reconstruction of these CT axons revealed that they are all collaterals of long-range corticofugal axons. These collaterals do not give off axonal branches within the thalamic reticular nucleus and they arborize exclusively within intralaminar and associative thalamic nuclei where they from small clusters of varicose endings. As layer V cells are involved in motor commands everywhere in the neocortex, these CT projections and their thalamic targets should be directly involved in the central organization of motor programs.

Intracellular current clamp recordings were obtained from mossy cells (n = 6, identified by intracellular injection of biocytin) of the dorsal dentate gyrus from rats under ketamine-xylazine anesthesia. During electroencephalographic theta rhythm (4-6 Hz), recorded with a macroelectrode placed in the contralateral dorsal hippocampus near the fissure, mossy cells displayed intracellular membrane potential oscillations at low frequencies (4-6 Hz) which appeared to be phase locked to the electroencephalographic theta rhythm. The frequency of the intracellular theta rhythm was independent of the membrane potential. However, the phase difference between the intracellular and the electroencephalographic theta rhythms as well as the amplitude of the intracellular theta oscillations were voltage-dependent. These findings are consistent with the hypothesis that rhythmic GABAA receptor-mediated inhibitory postsynaptic potentials contribute to the genesis of the intracellular theta rhythm. Indeed, mossy cells displayed an early, fast inhibitory postsynaptic potential in response to electrical stimulation of the entorhinal cortex, which most likely represents a GABAA receptor-mediated event, indicating that mossy cells possess functional GABAA receptors. At the resting membrane potential, mossy cells did not fire at each cycle of the electroencephalographic theta rhythm but fired only rarely (< 1 Hz). However, when they did fire they did so preferentially in phase with the peak positivity of the electroencephalographic theta rhythm. Reconstruction of two mossy cells with axonal projections to the inner molecular layer showed that the spatial extent of the influence such weakly discharging mossy cells may have on other dentate gyrus neurons during theta oscillations can be several millimeters in the septotemporal direction. In conclusion, these findings show that mossy cells of the rat hilus during ketamine-xylazine anesthesia participate in theta oscillations of the hippocampal formation, during which their low-frequency firing may contribute to the phase-locking of a large number of spatially distributed postsynaptic neurons with postsynaptic sites in the inner molecular layer of the dentate gyrus.

Formula are developed for calculating the performance characteristics (surface-to-total-volume ratio, surface-to-packing-volume ratio, and void volume fraction) of four different types of immobilized cell modular units (ICMUs) for ethanol fermentation: plate-type, spiral-type, beehive-type and bead-type ICMUs. Examples are used to illustrate how the formulas are useful for investigating the effects of characteristic dimensions of packing geometry, as well as the effect of scale on the performance characteristics of the ICMUs.