Addressing Electroconvulsive Therapy Knowledge Gaps and Stigmatized Views Among Nursing Students Through a Psychiatrist-APRN Didactic Partnership

BACKGROUND:Knowledge gaps and stigmatized perceptions regarding electroconvulsive therapy (ECT) among patients and health providers contribute to the underutilization of an important therapeutic modality. The proactive education of future advanced practice registered nurses (APRNs) provides an opportunity to optimize the use of this evidence-based clinical practice.AIMS:As part of a general course in psychiatry during the first year of nursing school, we dedicated 1 hour to treatment-refractory depression, including ECT, and a second hour to a summary discussion of mood disorders. We evaluated the efficacy of this didactic offering, which was co-taught by a psychiatrist and a psychiatric APRN.METHOD:At baseline, consenting students (n= 94) provided three words they associated with ECT and then completed three validated instruments: (a) Questionnaire on Attitudes and Knowledge of ECT, (b) Opening Minds Stigma Scale for Health Care Providers, and (c) Self-Stigma of Seeking Help. Among the 67 students who repeated the assessment at endpoint, 39 attended the ECT didactic (Intervention group, 58%) and 28 did not (Control, 42%).RESULTS:After completion of the 3-month course, students showed improvement across all measures (p<.001). The only outcomes that improved differentially between the Intervention and Control groups were the Questionnaire on Attitudes and Knowledge of ECT Attitudes and Knowledge scales (p= .01). Word choice valence associated with ECT shifted favorably by endpoint (p<.001).CONCLUSIONS:An educational intervention co-led by a psychiatric-mental health APRN had a significant impact on nursing students' knowledge and perceptions of ECT. This approach can be readily implemented at other institutions. Future refinements will include the videotaped depiction of a simulated patient undergoing the consent, treatment, and recovery phases of ECT

Dealing with Misfolded Proteins: Examining the Neuroprotective Role of Molecular Chaperones in Neurodegeneration

Human neurodegenerative diseases arise from a wide array of genetic and environmental factors. Despite the diversity in etiology, many of these diseases are considered "conformational" in nature, characterized by the accumulation of pathological, misfolded proteins. These misfolded proteins can induce cellular stress by overloading the proteolytic machinery, ultimately resulting in the accumulation and deposition of aggregated protein species that are cytotoxic. Misfolded proteins may also form aberrant, non-physiological protein-protein interactions leading to the sequestration of other normal proteins essential for cellular functions. The progression of such disease may therefore be viewed as a failure of normal protein homeostasis, a process that involves a network of molecules regulating the synthesis, folding, translocation and clearance of proteins. Molecular chaperones are highly conserved proteins involved in the folding of nascent proteins, and the repair of proteins that have lost their typical conformations. These functions have therefore made molecular chaperones an active area of investigation within the field of conformational diseases. This review will discuss the role of molecular chaperones in neurodegenerative diseases, highlighting their functional classification, regulation, and therapeutic potential for such diseases

518. Long-term lentiviral vector-mediated transgene expression in neural progenitor cells following implantation into the injured rat spinal cord

Due to their self-renewal and multi-potency, stem cells represent an attractive source for cell replacement therapy in neurological disorders. Genetic manipulation of these cells may allow controlled release of therapeutic proteins, suppress immune rejection, or produce essential neurotransmitters. Furthermore, when the expression cassette is incorporated into the host genome ex vivo, this technique also may be used as a method to trace cells following implantation into tissues of interest. We explored the possibility of transducing pluripotent fetal rat cortical neural progenitor cells using lentiviral vectors encoding either the green fluorescent protein (GFP) or neurotrophic factors (NT-3, BDNF, GDNF and CNTF) under control of the CMV promoter and the Woodchuck post-transcriptional regulatory element. Following isolation and expansion of the cells at clonal density on poly-ornithine-fibronectin-coated dishes in the presence of bFGF, cells were collected, infected and replated on the dishes. Staining of these cells for neural markers (such as nestin, GFAP, Tuj-1 and RIP) after transduction did not reveal any significant difference from non-transduced cells. However, when they were transduced with a vector encoding CNTF, cells started expressing GFAP. Cells continued to express the transgene, including when bFGF was withdrawn and when cells started to differentiate into GFAP positive cells. Following delayed (1 week) implantation into the lesion site of the moderately contused spinal cord, transduced cells survived well up to 4 weeks post-implantation (the longest time point currently examined). Migration of the cells was mainly restricted to white matter on either side of the lesion. Currently, the therapeutic and axonal growth stimulating properties of the implanted cells are being investigated in injured rats

662. The Tracking of Exogenous Cells in the Injured Central Nervous System: Viral Vector Labeling Versus an Intrinsic Genetic Marker

Transplantation of exogenous cells into the injured central nervous system and subsequent examination of how these cells survive, integrate and perform physiological functions within the host environment requires that the cells be clearly identifiable from host cells. As many of the cells used for transplantation studies are derived from cultures of cells indistinguishable from the host using immunochemical approaches, numerous dyes or genetic markers have been devised and used for this purpose. These markers used to label and track these transplanted cells need to be specific, reliable, resistant to leakage or cellular transfer, and remain chemically stable once inside the host to allow long-term tracking. In the current study we compared two labeling methods for long-term tracking of cells following transplantation into the injured rat spinal cord; lentiviral vector transduction with green fluorescent protein (GFP) and DNA in situ hybridization for the Y chromosome after male cell transplantation in female rats. Examination of labeled cells at 12 wk post-injury demonstrated that both labels did not exhibit significant fading or loss of the signal due to instability over time. The visualization procedure for the DNA in situ reaction product was significantly more labor intensive than simple identification of GFP by fluorescent microscopy. Y chromosome positive cells were, however, easier to definitely count for assessment of cell survival than GFP labeled cells, due to the presence of the label as a single point within the nucleus rather than diffuse labeling throughout the entirety of the cell. The limited presence of the signal in Y chromosome positive cells though, made examination of their ability to integrate within the injured spinal cord, migrate and associate with axons impossible without additional immunochemical methods for labeling the entire cell. The combination of immunochemistry with DNA in situ however proved to be unsuccessful with the many antibody combinations tested. In conclusion GFP delivery by lentiviral vectors was far superior to DNA in situ detection of the Y chromosome for labeling of transplanted cells due to its rapid visualization and its applicability to a wide range of assessment techniques

Lentiviral vector-mediated transduction of neural progenitor cells before implantation into injured spinal cord and brain to detect their migration, deliver neurotrophic factors and repair tissue

Stem cells represent an attractive source for cell replacement therapy in neurological disorders due to their self-renewal and multi-potency. Genetic manipulation of these cells may allow controlled release of therapeutic proteins, suppress immune rejection, or produce essential neurotransmitters. Furthermore, when the expression cassette is incorporated into the host genome ex vivo, this technique also may be used as a method to trace cells following implantation into tissues of interest. We explored the possibility of transducing pluripotent fetal rat cortical neural progenitor cells (NPCs) using lentiviral vectors encoding the green fluorescent protein (GFP) or neurotrophic factors (BDNF, CNTF, D15A, GDNF, MNT and NT-3) prior to implanting these cells into the contused spinal cord or injured brain. In vitro staining of these cells for neural markers (such as nestin, GFAP, Tuj-1 and RIP) after transduction did not reveal any significant difference from non-transduced cells. When they were transduced with a vector encoding CNTF or MNT, however, cells started expressing GFAP in vitro. Following delayed (1 week) implantation into the lesion site of the moderately contused rat spinal cord or the injured brain, transduced cells survived up to 12 weeks post-implantation (the longest time point examined) and most of the NPCs turned into an astrocytic phenotype in the spinal cord, but not in the brain. Nestin and GFP positive cells were detected in the brain, but not in the spinal cord lesion. GFP positive cells in the spinal cord migrated rostrally and caudally from the lesion/implantation site towards uninjured tissue. Novel findings in this study are the longterm expression of a foreign gene in NPCs using lentiviral vectors; this enabled tracking of the cells following implantation. This expression also allowed the observation that NPCs developed differently in the injured spinal cord and brain. Moreover, NPCs could be transduced to overexpress neurotrophic factors. In sum, NPC survival and the long-term transgene expression that allows easy tracking of migrating cells make NPCs promising candidates for implantation into the injured spinal cord or brain and a potentially powerful tool to enhance regeneration when transduced ex vivo to produce therapeutic molecules

Intramuscular AAV delivery of NT-3 alters synaptic transmission to motoneurons in adult rats

We examined whether elevating levels of neurotrophin-3 (NT-3) in the spinal cord and dorsal root ganglion (DRG) would alter connections made by muscle spindle afferent fibers on motoneurons. Adeno-associated virus (AAV) serotypes AAV1, AAV2 and AAV5, selected for their tropism profile, were engineered with the NT-3 gene and administered to the medial gastrocnemius muscle in adult rats. ELISA studies in muscle, DRG and spinal cord revealed that NT-3 concentration in all tissues peaked about 3 months after a single viral injection; after 6 months NT-3 concentration returned to normal values. Intracellular recording in triceps surae motoneurons revealed complex electrophysiological changes. Moderate elevation in cord NT-3 resulted in diminished segmental excitatory postsynaptic potential (EPSP) amplitude, perhaps as a result of the observed decrease in motoneuron input resistance. With further elevation in NT-3 expression, the decline in EPSP amplitude was reversed indicating that NT-3 at higher concentration could increase EPSP amplitude. No correlation was observed between EPSP amplitude and NT-3 concentration in the DRG. Treatment with control viruses could elevate NT-3 levels minimally resulting in measurable electrophysiological effects, perhaps as a result of inflammation associated with injection. EPSPs elicited by stimulation of the ventrolateral funiculus underwent a consistent decline in amplitude independent of NT-3 level. These novel correlations between modified NT-3 expression and single-cell electrophysiological parameters indicate that intramuscular administration of AAV(NT-3) can exert long lasting effects on synaptic transmission to motoneurons. This approach to neurotrophin delivery could be useful in modifying spinal function after injury

Transplantation of Schwann cells and/or olfactory ensheathing glia into the contused spinal cord: Survival, migration, axon association, and functional recovery

Schwann cells (SCs) and olfactory ensheathing glia (OEG) have shown promise for spinal cord injury repair. We sought their in vivo identification following transplantation into the contused adult rat spinal cord at 1 week post-injury by: (i) DNA in situ hybridization (ISH) with a Y-chromosome specific probe to identify male transplants in female rats and (ii) lentiviral vector-mediated expression of EGFP. Survival, migration, and axon-glia association were quantified from 3 days to 9 weeks post-transplantation. At 3 weeks after transplantation into the lesion, a 60-90% loss of grafted cells was observed. OEG-only grafts survived very poorly within the lesion (<5%); injection outside the lesion led to a 60% survival rate, implying that the injury milieu was hostile to transplanted cells and or prevented their proliferation. At later times post-grafting, p75(+)/EGFP(-) cells in the lesion outnumbered EGFP(+) cells in all paradigms, evidence of significant host SC infiltration. SCs and OEG injected into the injury failed to migrate from the lesion. Injection of OEG outside of the injury resulted in their migration into the SC-injected injury site, not via normal-appearing host tissue but along the pia or via the central canal. In all paradigms, host axons were seen in association with or ensheathed by transplanted glia. Numerous myelinated axons were found within regions of grafted SCs but not OEG. The current study details the temporal survival, migration, axon association of SCs and OEG, and functional recovery after grafting into the contused spinal cord, research previously complicated due to a lack of quality, long-term markers for cell tracking in vivo