Reduced axon sprouting after treatment that diminishes microglia accumulation at lesions in the leech CNS

Abstract

The role of mammalian microglia in central nervous system (CNS) repair is controversial. Microglia accumulate at lesions where they act as immune cells and phagocytize debris, and they may secrete neurotrophins, but they also produce molecules that can be cytotoxic, like nitric oxide (NO). To determine the importance of microglial accumulation at lesions on growth of severed CNS axons in the leech (Hirudo medicinalis), in which axon and synapse regeneration are notably successful even when isolated in tissue culture medium, microglial migration to lesions was reduced. Pressure (P) sensory neurons were injected with biocytin to reveal the extent of their sprouting 24 hours after lesioning. To reduce microglia accumulation at lesions, cords were treated for 3.5 hours with 3 mM ATP or 2 mM N(omega)-nitro-L-arginine methyl ester (L-NAME) or 50 microM Reactive blue-2 (RB2) beginning 30 minutes before injury. Lesioned controls were either not treated with drug or treated 3 hours later with one of the drugs, after the migration and subsequent accumulation of most microglia had occurred, but before the onset of axon sprouting, for a total of seven separate conditions. There was a significant reduction in total sprout lengths compared with controls when microglial accumulation was reduced. The results suggest that microglial cells are necessary for the usual sprouting of injured axons.

Physiology of Microglia

“…The accumulation of microglia is a prerequisite for axonal sprouting. When microglial accumulation was inhibited by 3 mM ATP or a NOS inhibitor, there was a significant reduction in total sprout lengths compared with controls when microglial accumulation was reduced (640). Thus, in the leech nervous system, microglial cells are important elements to control regeneration (631).…”

Nitric oxide regulates axonal regeneration in an insect embryonic CNS

“…The release of NO induces migration of microglia to the site of injury (Chen et al, 2000) in a cGMP-dependent manner (Duan et al, 2003). Preventing microglia migration by interfering with NO/cGMP signaling impairs regeneration (Ngu et al, 2007), indicating the importance of this mechanism for leech CNS regeneration.…”

“…Most invertebrates do not express myelin or its associated factors and lack CSPGs. Consequently, regeneration of both the peripheral and the central nervous system has been shown in several groups of invertebrates, such as snails (Moffett, 1995), leeches (Carbonetto and Muller, 1972;Duan et al, 2003, Ngu et al, 2007, or crayfish (Bittner, 1991). Among invertebrates, insects belong to the most evolved groups, displaying a high degree of complexity of their nervous systems.…”

Interaction of HmC1q with leech microglial cells: involvement of C1qBP-related molecule in the induction of cell chemotaxis

“…After injury, leech microglia immediately move toward the lesion site. This phenomenon has been shown to be essential to promote axon sprouting and successful nervous system repair [11-14]. Leech microglial cells exhibit morphological changes similar to vertebrate ones in the course of migration in response to tissue damage [15,16].…”