Age-related loss of VGLUT1 excitatory, but not VGAT inhibitory, immunoreactive terminals on motor neurons in spinal cords of old sarcopenic male mice

Abstract

Age-related changes in ventral lumbar spinal cord (L3-L5) were compared in young [3 month, (M)] and old (27 M) C57BL/6J male mice. The aged mice had previously been shown to exhibit sarcopenia and changes to peripheral nerve morphology. The putative connectivity of β-III tubulin positive α-motor neurons was compared in immunostained transverse sections using excitatory and inhibitory terminal markers vesicular glutamate transporter-1 (VGLUT1) and vesicular GABA transporter (VGAT). Glial fibrillary acidic protein (GFAP) and ionized calcium binding adaptor molecule 1 (Iba1) immunostaining was used to monitor changes in astrocyte and microglial phenotype respectively. For a given motor neuron, the neuronal perimeter was outlined and terminals immunoreactive for VGLUT1 or VGAT in close apposition to the soma were identified. By 27 M, the percentage coverage and total number of VGLUT1 immunoreactive terminals immediately adjacent to the soma of α-motor neurons was significantly decreased compared with young mice. However, percentage coverage of immunoreactive VGAT inhibitory terminals did not change significantly with age. The gray matter of 27 M spinal cords showed increased astrocytic and microglial activity. The loss of VGLUT1 terminals on α-motor neurons, terminals known to be derived from proprioceptive muscle afferents, may further impair sensorimotor control of hind limb skeletal muscle function in old mice.

Motoneuron deafferentation and gliosis occur in association with neuromuscular regressive changes during ageing in mice

“…Although MN death does not appear to be a relevant pathological process over the course of ageing, we cannot rule out the possibility that MNs with a normal histological appearance may have impaired function. In fact, we provide here strong evidence that MNs of old animals suffer a prominent loss of excitatory cholinergic and glutamatergic inputs, which is in agreement with recent reports 100,106 . Although we noticed no age‐associated changes in the density of inhibitory GABAergic synapses, their size on old MNs was found significantly reduced.…”

“…In fact, we provide here strong evidence that MNs of old animals suffer a prominent loss of excitatory cholinergic and glutamatergic inputs, which is in agreement with recent reports. 100,106 Although we noticed no age-associated changes in the density of inhibitory GABAergic synapses, their size on old MNs was found significantly reduced. MN deafferentation has also been widely reported by us and others in murine models of MN diseases, such as spinal muscular atrophy (SMA) [107][108][109][110][111] and amyotrophic lateral sclerosis (ALS), 109,[112][113][114][115] and also following nerve injury.…”

Dystrophic Dmd rats show early neuronal changes (increased S100β and Tau5) at 8 months, supporting severe dystropathology in this rodent model of Duchenne muscular dystrophy

“…It is emphasised that disturbances to the intact mature dystrophic (DMD) nerves are presumed to occur in response to altered neuronal connections at the level of NMJs in the distant target myofibres, as a result of repeated bouts of intrinsic necrosis in dystrophic skeletal muscles throughput the body (indicated in Figure 6) (Pannerec et al, 2016) and central changes to motor neurons in the spinal cord (Krishnan et al, 2018). These linked age-related events occur in a similar timeline (in the absence of any marked necrosis of healthy ageing muscles), although which components of the neuromuscular system drive sarcopenia remain unclear.…”

“…The confirmation of these neuronal changes in the 8M Dmd mdx rats, combined with data now for mdx mice aged 9M (as well as previous nerve data for 13M-18M old mdx mice), validates the use of these new neuronal markers as a useful indirect pre-clinical readout (using immunoblotting) to monitor the extent of ongoing myonecrosis in dystrophic muscles. Clearly more pre-clinical studies are required to provide detailed information, to clarify the reasons and implications, the earliest manifestation and later consequences of these neuronal changes, including the extent to which spinal cord connectivity (Krishnan et al, 2018) may show alterations at much older ages. It seems especially important to extend these neuronal studies to GRMD (and other dog) models of DMD where the disease manifestation is more pronounced: this reflects the relatively greater length of the growth phase (associated with pronounced myonecrosis), much bigger overall size (affecting muscle loading), and the much longer life-span of dystrophic dogs (allowing more time for manifestation of secondary progressive neuronal changes), compared with rodents.…”

Regeneration of adult rat sensory and motor neuron axons through chimeric peroneal nerve grafts containing donor Schwann cells engineered to express different neurotrophic factors

“…We examined immunostained sections from three normal, autograft, acellular and LV NT-3 graft animals, and from four SC-GFP, LV-CNTF and LV-NGF animals; in total, 370 FG + motor neurons were analyzed (Table 1). Only VGLUT-1 terminals that were in very close apposition to FG + cells were counted (Liu et al, 2014;Krishnan et al, 2018).…”