Health-system drivers influencing the continuum of care linkages for low-birth-weight infants at the different care levels in Ghana.

BACKGROUND: Low birth weight (LBW) is associated with short and long-term consequences including neonatal mortality and disability. Effective linkages in the continuum of care (CoC) for newborns at the health facility, community (primary care) and home care levels have a high tendency of minimizing adverse events associated with LBW. But it is unclear how these linkages work and what factors influence the CoC process in Ghana as literature is scarce on the views of health professionals and families of LBW infants regarding the CoC. Therefore, this study elicited the drivers influencing the CoC for LBW infants in Ghana and how linkages in the CoC could be strengthened to optimize quality of care. METHODS: A constructivist grounded theory study design was used. Data was collected between September 2020 to February 2021. A total of 25 interviews were conducted with 11 family members of LBW infants born in a secondary referral hospital in Ghana, 9 healthcare professionals and 7 healthcare managers. Audio recordings were transcribed verbatim, analyzed using initial and focused coding. Constant comparative techniques, theoretical memos, and diagramming were employed until theoretical saturation was determined. RESULTS: Emerging from the analysis was a theoretical model describing ten major themes along the care continuum for LBW infants, broadly categorized into health systems and family-systems drivers. In this paper, we focused on the former. Discharge, review, and referral systems were neither well-structured nor properly coordinated. Efficient dissemination and implementation of guidelines and supportive supervision contributed to higher staff motivation while insufficient investments and coordination of care activities limited training opportunities and human resource. A smooth transition between care levels is hampered by procedural, administrative, logistics, infrastructural and socio-economic barriers. CONCLUSION: A coordinated care process established on effective communication across different care levels, referral planning, staff supervision, decreased staff shuffling, routine in-service training, staff motivation and institutional commitment are necessary to achieve an effective care continuum for LBW infants and their families.

Structure-function analysis suggests that the photoreceptor LITE-1 is a light-activated ion channel.

Sensation of light is essential for all organisms. The eye-less nematode Caenorhabditis elegans detects UV and blue light to evoke escape behavior. The photosensor LITE-1 absorbs UV photons with an unusually high extinction coefficient, involving essential tryptophans. Here, we modeled the structure and dynamics of LITE-1 using AlphaFold2-multimer and molecular dynamics (MD) simulations and performed mutational and behavioral assays in C. elegans to characterize its function. LITE-1 resembles olfactory and gustatory receptors from insects, recently shown to be tetrameric ion channels. We identified residues required for channel gating, light absorption, and mechanisms of photo-oxidation, involving a likely binding site for the peroxiredoxin PRDX-2. Furthermore, we identified the binding pocket for a putative chromophore. Several residues lining this pocket have previously been established as essential for LITE-1 function. A newly identified critical cysteine pointing into the pocket represents a likely chromophore attachment site. We derived a model for how photon absorption, via a network of tryptophans and other aromatic amino acids, induces an excited state that is transferred to the chromophore. This evokes conformational changes in the protein, possibly leading to a state receptive to oxidation of cysteines and, jointly, to channel gating. Electrophysiological data support the idea that LITE-1 is a photon and H2O2-coincidence detector. Other proteins with similarity to LITE-1, specifically C. elegans GUR-3, likely use a similar mechanism for photon detection. Thus, a common protein fold and assembly, used for chemoreception in insects, possibly by binding of a particular compound, may have evolved into a light-activated ion channel.

Experiences of families and health professionals along the care continuum for low-birth weight neonates: A constructivist grounded theory study.

AIMS: To explore the experiences of health professionals and families concerning supporting low-birth weight (LBW) infants along the continuum of care (CoC) in Ghana with the goal to unveil new strategies to improve the quality of neonatal care. DESIGN: A constructivist grounded theory. METHOD: Simultaneous data collection and analysis among health professionals alongside families with LBW infants from September 2020 to April 2021. The study used constructivist grounded theory strategies for data collection and analysis. RESULTS: The analysis of 25 interviews resulted in a theoretical model describing 10 themes along the CoC for LBW infants, categorized into health and family systems drivers. In this paper, we focus on the latter. Early bonding and family involvement were empowering. Mothers needed assistance in balancing hope and confidence which enabled them to render special care to their LBW infants. Providing mothers with financial and domestic support as well as creating awareness on newborn health in communities were important. CONCLUSION: To achieve family involvement, a coordinated CoC must entail key players and be culturally inclusive. It must be applied at all levels in the CoC process in a non-linear faction. This can help LBW infants to thrive and to reach their full developmental potential. IMPACT: The theoretical model developed shows the importance of family involvement through family systems care for a comprehensive response in addressing needs of health professionals and families with LBW infants and bridging the fragmentations in the neonatal CoC in Ghana. Context-tailored research on family systems care in the neonatal period is necessary to achieve a quality CoC for LBW infants and their families. PATIENT OR PUBLIC CONTRIBUTION: Caregivers and providers participated by granting in-depth interviews. Care providers further contributed through their feedback on preliminary findings.

Adherence to appointments for gestational diabetes testing and experiences with two-hour postprandial glucose test: a mixed-methods study in Ghana.

BACKGROUND: Failure to attend scheduled appointments is a common problem in healthcare. In obstetrics, diagnostic and treatment protocols for gestational diabetes mellitus (GDM) require client booking, test preparations, management and follow-up reviews. We identified the socio-demographic, obstetric and medical drivers influencing adherence to appointments for GDM testing and experiences of pregnant women's regarding performing oral glucose tolerance test (OGTT). METHODS: A convergent parallel mixed-methods study comprising a cross-sectional survey and an explorative qualitative descriptive design were used. We recruited 817 women in their first trimester of pregnancy from the antenatal clinics of primary, secondary and tertiary health facilities in Ghana. After obtaining their demographic and health history, we scheduled them for 2-h OGTT between 24 and 28 gestational weeks and estimated the odds of returning for the test. In the qualitative phase, we called 166 participants to ascertain why they failed to report. Also, we had in-depth and focused group discussions with 60 postpartum women who performed the OGTT to explore their experiences with the test. RESULTS: Out of 817 pregnant women scheduled, 490 (59.97%) reported of which 54.59, 54.33 and 53.24% completed fasting plasma glucose, 1-h and 2-h OGTT, respectively. Maternal age above 35 years (OR: 3.56, 95% CI:1.49-8.47), secondary education (OR: 3.21, 95% CI: 1.19-8.69), formal sector employment (OR: 2.02, 95% CI: 1.16-3.51) and having same-sex children (OR: 4.37, 95% CI: 1.98-9.66) increased odds of appointment adherence whereas healthcare in a tertiary hospital (OR:0.46, 95% CI:0.22-0.96), rural residence (OR: 0.53, 95% CI: 0.34-0.85) and being overweight (OR: 0.45, 95% CI: 0.25-0.78) decreased the likelihood. Experiences were thematized into feelings about test procedure, acceptability of test, skillfulness of the health workers and information on the test. Despite the apprehension and discomforts associated with the test, the desire to know one's disease status was the chief motivation. Empathy, reassurance and receiving ample information on the test procedures eased anxiety and improved test compliance. CONCLUSIONS: Although 40% of participants scheduled did not return, the test was generally acceptable. Socio-cultural underpinnings influenced the health-seeking behaviors, meaning that health worker interactions on test procedures need to be sensitive to the woman's situation.

An Optogenetic Arrhythmia Model-Insertion of Several Catecholaminergic Polymorphic Ventricular Tachycardia Mutations Into Caenorhabditis elegans UNC-68 Disturbs Calstabin-Mediated Stabilization of the Ryanodine Receptor Homolog.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited disturbance of the heart rhythm (arrhythmia) that is induced by stress or that occurs during exercise. Most mutations that have been linked to CPVT are found in two genes, i.e., ryanodine receptor 2 (RyR2) and calsequestrin 2 (CASQ2), two proteins fundamentally involved in the regulation of intracellular Ca2+ in cardiac myocytes. We inserted six CPVT-causing mutations via clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 into unc-68 and csq-1, the Caenorhabditis elegans homologs of RyR and CASQ, respectively. We characterized those mutations via video-microscopy, electrophysiology, and calcium imaging in our previously established optogenetic arrhythmia model. In this study, we additionally enabled high(er) throughput recordings of intact animals by combining optogenetic stimulation with a microfluidic chip system. Whereas only minor/no pump deficiency of the pharynx was observed at baseline, three mutations of UNC-68 (S2378L, P2460S, Q4623R; RyR2-S2246L, -P2328S, -Q4201R) reduced the ability of the organ to follow 4 Hz optogenetic stimulation. One mutation (Q4623R) was accompanied by a strong reduction of maximal pump rate. In addition, S2378L and Q4623R evoked an altered calcium handling during optogenetic stimulation. The 1,4-benzothiazepine S107, which is suggested to stabilize RyR2 channels by enhancing the binding of calstabin2, reversed the reduction of pumping ability in a mutation-specific fashion. However, this depends on the presence of FKB-2, a C. elegans calstabin2 homolog, indicating the involvement of calstabin2 in the disease-causing mechanisms of the respective mutations. In conclusion, we showed for three CPVT-like mutations in C. elegans RyR a reduced pumping ability upon light stimulation, i.e., an arrhythmia-like phenotype, that can be reversed in two cases by the benzothiazepine S107 and that depends on stabilization via FKB-2. The genetically amenable nematode in combination with optogenetics and high(er) throughput recordings is a promising straightforward system for the investigation of RyR mutations and the selection of mutation-specific drugs.

A GABAergic and peptidergic sleep neuron as a locomotion stop neuron with compartmentalized Ca2+ dynamics.

Animals must slow or halt locomotion to integrate sensory inputs or to change direction. In Caenorhabditis elegans, the GABAergic and peptidergic neuron RIS mediates developmentally timed quiescence. Here, we show RIS functions additionally as a locomotion stop neuron. RIS optogenetic stimulation caused acute and persistent inhibition of locomotion and pharyngeal pumping, phenotypes requiring FLP-11 neuropeptides and GABA. RIS photoactivation allows the animal to maintain its body posture by sustaining muscle tone, yet inactivating motor neuron oscillatory activity. During locomotion, RIS axonal Ca2+ signals revealed functional compartmentalization: Activity in the nerve ring process correlated with locomotion stop, while activity in a branch correlated with induced reversals. GABA was required to induce, and FLP-11 neuropeptides were required to sustain locomotion stop. RIS attenuates neuronal activity and inhibits movement, possibly enabling sensory integration and decision making, and exemplifies dual use of one cell across development in a compact nervous system.

Rhodopsin-based voltage imaging tools for use in muscles and neurons of Caenorhabditis elegans.

Genetically encoded voltage indicators (GEVIs) based on microbial rhodopsins utilize the voltage-sensitive fluorescence of all-trans retinal (ATR), while in electrochromic FRET (eFRET) sensors, donor fluorescence drops when the rhodopsin acts as depolarization-sensitive acceptor. In recent years, such tools have become widely used in mammalian cells but are less commonly used in invertebrate systems, mostly due to low fluorescence yields. We systematically assessed Arch(D95N), Archon, QuasAr, and the eFRET sensors MacQ-mCitrine and QuasAr-mOrange, in the nematode Caenorhabditis elegans ATR-bearing rhodopsins reported on voltage changes in body wall muscles (BWMs), in the pharynx, the feeding organ [where Arch(D95N) showed approximately 128% ΔF/F increase per 100 mV], and in neurons, integrating circuit activity. ATR fluorescence is very dim, yet, using the retinal analog dimethylaminoretinal, it was boosted 250-fold. eFRET sensors provided sensitivities of 45 to 78% ΔF/F per 100 mV, induced by BWM action potentials, and in pharyngeal muscle, measured in simultaneous optical and sharp electrode recordings, MacQ-mCitrine showed approximately 20% ΔF/F per 100 mV. All sensors reported differences in muscle depolarization induced by a voltage-gated Ca2+-channel mutant. Optogenetically evoked de- or hyperpolarization of motor neurons increased or eliminated action potential activity and caused a rise or drop in BWM sensor fluorescence. Finally, we analyzed voltage dynamics across the entire pharynx, showing uniform depolarization but compartmentalized repolarization of anterior and posterior parts. Our work establishes all-optical, noninvasive electrophysiology in live, intact C. elegans.

Mothers' Experiences with Neonatal Care for Low Birth Weight Infants at Home; A Qualitative Study in the Hohoe Municipality, Ghana.

PURPOSE: To explore knowledge and beliefs of mothers on low birth weight (LBW), examine care provision at home and societal perceptions of LBW infants. DESIGN AND METHODS: This qualitative study was conducted using hermeneutic phenomenological approach. Data of mothers who delivered LBW infants within 2 years preceding the study were purposively extracted from the medical records of the Hohoe Municipality Hospital in Ghana. Twenty semi-structured interviews and three focus group discussions were conducted. A thematic analysis approach was performed using Atlas.ti. RESULTS: Mothers identified and described LBW babies based on frailty, size and activity levels. LBW recognition was easier for multiparous mothers by comparing with previous deliveries. LBW was linked to poor maternal diet, diseases during pregnancy and heavy workload. Although most mothers perceived their LBW babies as healthy irrespective of the size a few home-care practises differed. Smaller LBW infants were less likely to be socially accepted. In the first few weeks after birth the care of LBW infants is the core responsibility of grandmothers. Primiparous mothers and those whose infants were smaller (<2 kg) quested for more information and support on LBW newborn care at home. CONCLUSION: There is a need to increase knowledge on risk factors and tackle lapses in the recognition and care of LBW infants. Counselling on recommended neonatal care should begin during antenatal care and reiterated during postnatal care. PRACTICAL IMPLICATION: Tailored in-depth and culturally-adapted counselling, discharge instructions and home-based postnatal visits targeted at LBW infants and their primary caregivers could improve care.

An optogenetic arrhythmia model to study catecholaminergic polymorphic ventricular tachycardia mutations.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a condition of abnormal heart rhythm (arrhythmia), induced by physical activity or stress. Mutations in ryanodine receptor 2 (RyR2), a Ca2+ release channel located in the sarcoplasmic reticulum (SR), or calsequestrin 2 (CASQ2), a SR Ca2+ binding protein, are linked to CPVT. For specific drug development and to study distinct arrhythmias, simple models are required to implement and analyze such mutations. Here, we introduced CPVT inducing mutations into the pharynx of Caenorhabditis elegans, which we previously established as an optogenetically paced heart model. By electrophysiology and video-microscopy, we characterized mutations in csq-1 (CASQ2 homologue) and unc-68 (RyR2 homologue). csq-1 deletion impaired pharynx function and caused missed pumps during 3.7 Hz pacing. Deletion mutants of unc-68, and in particular the point mutant UNC-68(R4743C), analogous to the established human CPVT mutant RyR2(R4497C), were unable to follow 3.7 Hz pacing, with progressive defects during long stimulus trains. The pharynx either locked in pumping at half the pacing frequency or stopped pumping altogether, possibly due to UNC-68 leakiness and/or malfunctional SR Ca2+ homeostasis. Last, we could reverse this 'worm arrhythmia' by the benzothiazepine S107, establishing the nematode pharynx for studying specific CPVT mutations and for drug screening.

Arrhythmogenic effects of mutated L-type Ca 2+-channels on an optogenetically paced muscular pump in Caenorhabditis elegans.

Cardiac arrhythmias are often associated with mutations in ion channels or other proteins. To enable drug development for distinct arrhythmias, model systems are required that allow implementing patient-specific mutations. We assessed a muscular pump in Caenorhabditis elegans. The pharynx utilizes homologues of most of the ion channels, pumps and transporters defining human cardiac physiology. To yield precise rhythmicity, we optically paced the pharynx using channelrhodopsin-2. We assessed pharynx pumping by extracellular recordings (electropharyngeograms--EPGs), and by a novel video-microscopy based method we developed, which allows analyzing multiple animals simultaneously. Mutations in the L-type VGCC (voltage-gated Ca(2+)-channel) EGL-19 caused prolonged pump duration, as found for analogous mutations in the Cav1.2 channel, associated with long QT syndrome. egl-19 mutations affected ability to pump at high frequency and induced arrhythmicity. The pharyngeal neurons did not influence these effects. We tested whether drugs could ameliorate arrhythmia in the optogenetically paced pharynx. The dihydropyridine analog Nemadipine A prolonged pump duration in wild type, and reduced or prolonged pump duration of distinct egl-19 alleles, thus indicating allele-specific effects. In sum, our model may allow screening of drug candidates affecting specific VGCCs mutations, and permit to better understand the effects of distinct mutations on a macroscopic level.

Genetically encoded calcium indicators for multi-color neural activity imaging and combination with optogenetics.

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, "RCaMPs," engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca(2+)-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca(2+)]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca(2+) affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics.

Transport activity of the high-affinity monocarboxylate transporter MCT2 is enhanced by extracellular carbonic anhydrase IV but not by intracellular carbonic anhydrase II.

The ubiquitous enzyme carbonic anhydrase isoform II (CAII) has been shown to enhance transport activity of the proton-coupled monocarboxylate transporters MCT1 and MCT4 in a non-catalytic manner. In this study, we investigated the role of cytosolic CAII and of the extracellular, membrane-bound CA isoform IV (CAIV) on the lactate transport activity of the high-affinity monocarboxylate transporter MCT2, heterologously expressed in Xenopus oocytes. In contrast to MCT1 and MCT4, transport activity of MCT2 was not altered by CAII. However, coexpression of CAIV with MCT2 resulted in a significant increase in MCT2 transport activity when the transporter was coexpressed with its associated ancillary protein GP70 (embigin). The CAIV-mediated augmentation of MCT2 activity was independent of the catalytic activity of the enzyme, as application of the CA-inhibitor ethoxyzolamide or coexpressing the catalytically inactive mutant CAIV-V165Y did not suppress CAIV-mediated augmentation of MCT2 transport activity. Furthermore, exchange of His-88, mediating an intramolecular H(+)-shuttle in CAIV, to alanine resulted only in a slight decrease in CAIV-mediated augmentation of MCT2 activity. The data suggest that extracellular membrane-bound CAIV, but not cytosolic CAII, augments transport activity of MCT2 in a non-catalytic manner, possibly by facilitating a proton pathway other than His-88.

Intramolecular proton shuttle supports not only catalytic but also noncatalytic function of carbonic anhydrase II.

Carbonic anhydrases (CAs) catalyze the reversible hydration of CO(2) to HCO(3)(-) and H(+). The rate-limiting step in this reaction is the shuttle of protons between the catalytic center of the enzyme and the bulk solution. In carbonic anhydrase II (CAII), the fastest and most wide-spread isoform, this H(+) shuttle is facilitated by the side chain of His64, whereas CA isoforms such as carbonic anhydrase III (CAIII), which lack such a shuttle, have only low catalytic activity in vitro. By using heterologous protein expression in Xenopus oocytes, we tested the role of this intramolecular H(+) shuttle on CA activity in an intact cell. The data revealed that CAIII, shown in vitro to have ∼1,000-fold reduced activity as compared with CAII, displays significant catalytic activity in the intact cell. Furthermore, we tested the hypothesis that the H(+) shuttle in CAII itself can facilitate transport activity of the monocarboxylate transporters 1 and 4 (MCT1/4) independent of catalytic activity. Our results show that His64 is essential for the enhancement of lactate transport via MCT1/4, because a mutation of this residue to alanine (CAII-H64A) abolishes the CAII-induced increase in MCT1/4 activity. However, injection of 4-methylimidazole, which acts as an exogenous H(+) donor/acceptor, can restore the ability of CAII-H64A to enhance transport activity of MCT1/4. These findings support the hypothesis that the H(+) shuttle in CAII not only facilitates CAII catalytic activity but also can enhance activity of acid-/base-transporting proteins such as MCT1/4 in a direct, noncatalytic manner, possibly by acting as an "H(+)-collecting antenna."