Nucleolar integrity is required for the maintenance of long-term synaptic plasticity.

Long-term memory (LTM) formation requires new protein synthesis and new gene expression. Based on our work in Aplysia, we hypothesized that the rRNA genes, stimulation-dependent targets of the enzyme Poly(ADP-ribose) polymerase-1 (PARP-1), are primary effectors of the activity-dependent changes in synaptic function that maintain synaptic plasticity and memory. Using electrophysiology, immunohistochemistry, pharmacology and molecular biology techniques, we show here, for the first time, that the maintenance of forskolin-induced late-phase long-term potentiation (L-LTP) in mouse hippocampal slices requires nucleolar integrity and the expression of new rRNAs. The activity-dependent upregulation of rRNA, as well as L-LTP expression, are poly(ADP-ribosyl)ation (PAR) dependent and accompanied by an increase in nuclear PARP-1 and Poly(ADP) ribose molecules (pADPr) after forskolin stimulation. The upregulation of PARP-1 and pADPr is regulated by Protein kinase A (PKA) and extracellular signal-regulated kinase (ERK)--two kinases strongly associated with long-term plasticity and learning and memory. Selective inhibition of RNA Polymerase I (Pol I), responsible for the synthesis of precursor rRNA, results in the segmentation of nucleoli, the exclusion of PARP-1 from functional nucleolar compartments and disrupted L-LTP maintenance. Taken as a whole, these results suggest that new rRNAs (28S, 18S, and 5.8S ribosomal components)--hence, new ribosomes and nucleoli integrity--are required for the maintenance of long-term synaptic plasticity. This provides a mechanistic link between stimulation-dependent gene expression and the new protein synthesis known to be required for memory consolidation.

Cellular and subcellular localization of PKMζ.

In contrast to protein kinases that participate in long-term potentiation (LTP) induction and memory consolidation, the autonomously active atypical protein kinase C isoform, protein kinase Mzeta (PKMζ), functions in the core molecular mechanism of LTP maintenance and long-term memory storage. Here, using multiple complementary techniques for light and electron microscopic immunolocalization, we present the first detailed characterization of the cellular and subcellular distribution of PKMζ in rat hippocampus and neocortex. We find that PKMζ is widely expressed in forebrain with prominent immunostaining in hippocampal and neocortical grey matter, and weak label in white matter. In hippocampal and cortical pyramidal cells, PKMζ expression is predominantly somatodendritic, and electron microscopy highlights the kinase at postsynaptic densities and in clusters within spines. In addition, nuclear label and striking punctate immunopositive structures in a paranuclear and dendritic distribution are seen by confocal microscopy, occasionally at dendritic bifurcations. PKMζ immunoreactive granules are observed by electron microscopy in cell bodies and dendrites, including endoplasmic reticulum. The widespread distribution of PKMζ in nuclei, nucleoli and endoplasmic reticulum suggests potential roles of this kinase in cell-wide mechanisms involving gene expression, biogenesis of ribosomes and new protein synthesis. The localization of PKMζ within postsynaptic densities and spines suggests sites where the kinase stores information during LTP maintenance and long-term memory.

Study on a nonhealing fracture from a patient with systemic lupus erythematosus and its pathogenetic mechanisms.

Arthritis and osteonecrosis affect a large number of patients with systemic lupus erythematosus (SLE). A patient with history of SLE suffered a traumatic fracture of the left foot. Despite a long period of immobilization and internal fixation, the fracture failed to heal and required arthrodeses with removal of the phalanx. Histopathological investigation revealed destruction of cartilage, subchondral cystic degeneration, vasculitis, deposition of fibrinogen, type III collagen and fibronectin, absence of bone remolding, and detectable F-actin. The nonhealing was therefore due to lack of progression of healing process beyond the initial stage. There was deposition of immunoglobulins and complement C4b, possibly forming immune complex by autoantibodies and cellular components. The authors found that MSE55 protein, required for polymerization of actin and initiation of cellular process organization, had a similar cellular deposition as that of immunoglobulins. Autoantibodies thus may inhibit differentiation of the bone cells, and resulted in nonunion in the patient.

Identification of Prevotella in pedal osteomyelitis of a diabetic patient.

Osteomyelitis in a diabetic patient with a nonhealing foot ulcer, multiple medical conditions, and recurrent hospitalization for antibiotic therapy was found to be associated with gram-negative bacteria Prevotella melanginoganica and Prevotella melaninoganica hemagglutinating variant. Those organisms were identified due to the morphologically distinct features in electron microscopy and sequencing of the genes after Polymerase chain reaction amplification from the pathological material. The bacteria invaded the bone and resided in osteocyte, osteoblast, and endothelial cells. The bacteria are usually associated with periodontal plaques, causing inflammation and destruction of gingival tissue and resorption of the alveolar bone. This is the first ultrastructural and molecular study of a diabetic bone lesion with anaerobic bacterial infection.