Endogenous opioid peptides contribute to associative LTP in the hippocampal CA3 region.

The medial and lateral perforant path projections to the hippocampal CA3 region display distinct mechanisms of long-term potentiation (LTP) induction, N-methyl-d-aspartate (NMDA) and opioid receptor dependent, respectively. However, medial and lateral perforant path projections to the CA3 region display associative LTP with coactivation, suggesting that while they differ in receptors involved in LTP induction they may share common downstream mechanisms of LTP induction. Here we address this interaction of LTP induction mechanisms by evaluating the contribution of opioid receptors to the induction of associative LTP among the medial and lateral perforant path projections to the CA3 region in vivo. Local application of the opioid receptor antagonists naloxone or Cys2-Tyr3-Orn5-Pen7-amide (CTOP) normally block induction of lateral perforant path-CA3 LTP. However, these opioid receptor antagonists failed to block associative LTP in lateral perforant path-CA3 synapses when it was induced by strong coactivation of the medial perforant pathway which displays NMDAR-dependent LTP. Thus strong activation of non-opioidergic afferents can substitute for the opioid receptor activation required for lateral perforant path LTP induction. Conversely, medial perforant path-CA3 associative LTP was blocked by opioid receptor antagonists when induced by strong coactivation of the opioidergic lateral perforant path. These data indicate endogenous opioid peptides contribute to associative LTP at coactive synapses when induced by strong coactivation of an opioidergic afferent system. These data further suggest that associative LTP induction is regulated by the receptor mechanisms of the strongly stimulated pathway. Thus, while medial and lateral perforant path synapses differ in their mechanisms of LTP induction, associative LTP at these synapses share common downstream mechanisms of induction.

Regulation of skeletal muscle regeneration by CCR2-activating chemokines is directly related to macrophage recruitment.

Muscle regeneration requires CC chemokine receptor 2 (CCR2) expression on bone marrow-derived cells; macrophages are a prominent CCR2-expressing cell in this process. CCR2-/- mice have severe impairments in angiogenesis, macrophage recruitment, and skeletal muscle regeneration following cardiotoxin (CTX)-induced injury. However, multiple chemokines activate CCR2, including monocyte chemotactic proteins (MCP)-1, -3, and -5. We hypothesized that MCP-1 is the chemokine ligand that mediates the impairments present in CCR2-/- mice. We examined muscle regeneration, capillary density, and cellular recruitment in MCP-1-/- and CCR2-/- mice following injury. Muscle regeneration and adipocyte accumulation, but not capillary density, were significantly impaired in MCP-1-/- compared with wild-type (WT) mice; however, muscle regeneration and adipocyte accumulation impairments were not as severe as observed in CCR2-/- mice. Although tissue levels of MCP-5 were elevated in MCP-1-/- mice compared with WT, the administration of MCP-5 neutralizing antibody did not alter muscle regeneration in MCP-1-/- mice. While neutrophil accumulation after injury was similar in all three mouse strains, macrophage recruitment was highest in WT mice, intermediate in MCP-1-/- mice, and severely impaired in CCR2-/- mice. In conclusion, while the absence of MCP-1 resulted in impaired macrophage recruitment and muscle regeneration, MCP-1-/- mice exhibit an intermediate phenotype compared with CCR2-/- mice. Intermediate macrophage recruitment in MCP-1-/- mice was associated with similar capillary density to WT, suggesting that fewer macrophages may be needed to restore angiogenesis vs. muscle regeneration. Finally, other chemokines, in addition to MCP-1 and MCP-5, may activate CCR2-dependent regenerative processes resulting in an intermediate phenotype in MCP-1-/- mice.

Crimson carrier, a long-acting contrast agent for in vivo near-infrared imaging of injured and diseased muscle.

The near-infrared wavelengths (700-900 nm) are the most suitable optical window for light penetration and deep tissue imaging in small animals. Herein we report a near-infrared fluorescent contrast agent, crimson carrier, which acts as a blood pool contrast agent to detect and quantify injury and disease in live animals. After determining the excitation-emission spectra and pharmacokinetics, crimson carrier was injected into myoinjured mice to monitor their recovery. Crimson carrier was also used to image transgenic mice with spontaneous tumors. Crimson carrier has maximal excitation and emission wavelengths of 745 nm and 820 nm, respectively. Elimination occurs predominantly via urinary excretion. We demonstrate the utility of this contrast agent for serial imaging of traumatized muscle as well as muscle tumors. The unique long-acting pharmacokinetics and urinary excretion route characteristics make crimson carrier a contrast agent of choice for the visualization of tumors and injured muscle or other tissues in live animal studies.

Near-infrared imaging of injured tissue in living subjects using IR-820.

The unprecedented increase in preclinical studies necessitates high-throughput, inexpensive, and straightforward methods for evaluating diseased tissues. Near-infrared imaging of live subjects is a versatile, cost-effective technology that can be effectively used in a variety of pathologic conditions. We have characterized an inexpensive optoelectronic chemical, IR-820, as an infrared blood pool contrast agent to detect and quantify diseased tissue in live animals. IR-820 has maximal excitation and emission wavelengths of 710 and 820 nm, respectively. IR-820 emission is significantly improved in vivo on serum binding to albumin, and elimination occurs predominantly via the gastrointestinal tract. We demonstrate the utility of this contrast agent for serially imaging of traumatized tissue (muscle), tissue following reperfusion (eg, stroke), and tumors. IR-820 can also be employed to map regional lymph nodes. This novel contrast agent is anticipated to be a useful and an inexpensive tool for screening a wide variety of preclinical models of human diseases.

Bone marrow-derived cell regulation of skeletal muscle regeneration.

Limb regeneration requires the coordination of multiple stem cell populations to recapitulate the process of tissue formation. Therefore, bone marrow (BM) -derived cell regulation of skeletal muscle regeneration was examined in mice lacking the CC chemokine receptor 2 (CCR2). Myofiber size, numbers of myogenic progenitor cells (MPCs), and recruitment of BM-derived cells and macrophages were assessed after cardiotoxin-induced injury of chimeric mice produced by transplanting BM from wild-type (WT) or CCR2(-/-) mice into irradiated WT or CCR2(-/-) host mice. Regardless of the host genotype, muscle regeneration and recruitment of BM-derived cells and macrophages were similar in mice replenished with WT BM, whereas BM-derived cells and macrophage accumulation were decreased and muscle regeneration was impaired in all animals receiving CCR2(-/-) BM. Furthermore, numbers of MPCs (CD34(+)/Sca-1(-)/CD45(-) cells) were significantly increased in mice receiving CCR2(-/-) BM despite the decreased size of regenerated myofibers. Thus, the expression of CCR2 on BM-derived cells regulated macrophage recruitment into injured muscle, numbers of MPC, and the extent of regenerated myofiber size, all of which were independent of CCR2 expression on host-derived cells. Future studies in regenerative medicine must include consideration of the role of BM-derived cells, possibly macrophages, in CCR2-dependent events that regulate effective skeletal muscle regeneration.

Airway bypass improves the mechanical properties of explanted emphysematous lungs.

RATIONALE: By creating artificial communications through bronchial walls into the parenchyma of explanted lungs (airway bypass), we expect to decrease the amount of gas trapped and to increase the rate and volume of air expelled during forced expirations. OBJECTIVES: To describe the mechanism by which airway bypass improves the mechanical properties of the emphysematous lung. METHODS: Lung compartments and mechanics were measured before and after airway bypass, which was created by placement of three or four stent-supported fenestrations in 10 emphysematous lungs removed at transplantation surgery. MEASUREMENTS AND MAIN RESULTS: Minimal volume after passive deflation decreased by a mean of 1.54 L (range, 0.7-2.5 L) or 60% (range, 37-86%). Explanted VC increased by 1.30 L or 132% (range, 78-318%). Maximal expiratory flows and volumes increased. Flow resistance decreased. CONCLUSIONS: Because these data show that airway bypass improves the mechanics of breathing in severely emphysematous lungs in vitro, there is now strong empirical support that this procedure can improve ventilatory function in patients by reducing gas trapping and flow resistance.

Associative long-term potentiation (LTP) among extrinsic afferents of the hippocampal CA3 region in vivo.

Monosynaptic perforant path projections to the CA3 region of the hippocampus are anatomically and physiologically substantial pathways that relay cortical input directly to the hippocampus proper. Despite the suggested relevance of these direct pathways in models of information processing within the CA3 region, surprisingly few studies have characterized synaptic plasticity in these direct cortical projections to the CA3 region. We assessed the ability of perforant path projections, and commissural/associational projections to the hippocampal CA3 region to both induce or display associative LTP in vivo. In pentobarbital-anesthetized adult rats, trains delivered to either the medial or lateral perforant pathway at current intensities normally insufficient to induce LTP displayed associative LTP when these same trains were delivered in conjunction with high-intensity trains to the alternate perforant pathway. Similarly, associative LTP is induced at intrinsic commissural/associational-CA3 (C/A-CA3) synapses when weak C/A trains were delivered in conjunction with high-intensity trains to either the medial or lateral perforant pathway. Associative LTP also was observed at medial and lateral perforant path-CA3 synapses when weak perforant path trains were tetanized in conjunction with high-intensity trains delivered to C/A-CA3 synapses. Thus direct perforant path-CA3 synapses and commissural/associational-CA3 synapses can modify and be modified by other CA3 afferents in an associative manner, verifying a requirement for synaptic plasticity explicit in models of autoassociative information processing in the CA3 region.

Long-term potentiation in direct perforant path projections to the hippocampal CA3 region in vivo.

The perforant path constitutes the primary projection system relaying information from the neocortex to the hippocampal formation. Long-term synaptic potentiation (LTP) in the perforant path projections to the dentate gyrus is well characterized. However, surprisingly few studies have addressed the mechanisms underlying LTP induction in the direct perforant path projections to the hippocampus. Here we investigate the role of N-methyl-D-aspartate (NMDA) and opioid receptors in the induction of LTP in monosynaptic medial and lateral perforant path projections to the CA3 region in adult pentobarbital sodium-anesthetized rats. Similar to LTP observed at the medial perforant path-dentate gyrus synapse, medial perforant path-CA3 synapses display LTP that is blocked by both local and systemic administration of the competitive NMDA receptor antagonist (+/-)-3-(2-carboxypiperazin-4-yl) propyl-1-phosphonic acid [(+/-)-CPP]. By contrast, LTP induced at the lateral perforant path-CA3 synapses is not blocked by either local or systemic administration of this NMDA receptor antagonist. The induction of LTP at lateral perforant path-CA3 synapses, which is blocked by the opioid receptor antagonist naloxone, is also blocked by the selective mu opioid receptor antagonist Cys(2)-Tyr(3)-Orn(5)-Pen(7)-amide (CTOP), but not the selective delta opioid receptor antagonist naltrindole (NTI). CTOP was without effect on the induction of medial perforant path-CA3 LTP. The selective sensitivity of lateral perforant path-CA3 LTP to mu-opioid receptor antagonists corresponds with the distribution of mu-opioid receptors within the stratum lacunosum-moleculare of area CA3 where perforant path projections to CA3 terminate. These data indicate that both lateral and medial perforant path projections to the CA3 region display LTP, and that LTP induction in medial and lateral perforant path-CA3 synapses are differentially sensitive to NMDA receptor and mu-opioid receptor antagonists. This suggests a role for opioid, but not NMDA receptors in the induction of LTP at lateral perforant path projections to the hippocampal formation.