Limits...
Visualization of Sensory Neurons and Their Projections in an Upper Motor Neuron Reporter Line.

Genç B, Lagrimas AK, Kuru P, Hess R, Tu MW, Menichella DM, Miller RJ, Paller AS, Özdinler PH - PLoS ONE (2015)

Bottom Line: Our findings corroborate previous studies, and suggest that more than five months is required for significant skin denervation.Crossing UCHL1-eGFP with hSOD1G93A mice generated hSOD1G93A-UeGFP reporter line of amyotrophic lateral sclerosis, and revealed sensory nervous system defects, especially towards disease end-stage.Our studies not only emphasize the complexity of the disease in ALS, but also reveal that UCHL1-eGFP reporter line would be a valuable tool to visualize and study various aspects of sensory nervous system development and degeneration in the context of numerous diseases.

View Article: PubMed Central - PubMed

Affiliation: Davee Department of Neurology and Clinical Neurological Sciences, Northwestern University, Feinberg School of Medicine, Chicago, IL, United States of America.

ABSTRACT
Visualization of peripheral nervous system axons and cell bodies is important to understand their development, target recognition, and integration into complex circuitries. Numerous studies have used protein gene product (PGP) 9.5 [a.k.a. ubiquitin carboxy-terminal hydrolase L1 (UCHL1)] expression as a marker to label sensory neurons and their axons. Enhanced green fluorescent protein (eGFP) expression, under the control of UCHL1 promoter, is stable and long lasting in the UCHL1-eGFP reporter line. In addition to the genetic labeling of corticospinal motor neurons in the motor cortex and degeneration-resistant spinal motor neurons in the spinal cord, here we report that neurons of the peripheral nervous system are also fluorescently labeled in the UCHL1-eGFP reporter line. eGFP expression is turned on at embryonic ages and lasts through adulthood, allowing detailed studies of cell bodies, axons and target innervation patterns of all sensory neurons in vivo. In addition, visualization of both the sensory and the motor neurons in the same animal offers many advantages. In this report, we used UCHL1-eGFP reporter line in two different disease paradigms: diabetes and motor neuron disease. eGFP expression in sensory axons helped determine changes in epidermal nerve fiber density in a high-fat diet induced diabetes model. Our findings corroborate previous studies, and suggest that more than five months is required for significant skin denervation. Crossing UCHL1-eGFP with hSOD1G93A mice generated hSOD1G93A-UeGFP reporter line of amyotrophic lateral sclerosis, and revealed sensory nervous system defects, especially towards disease end-stage. Our studies not only emphasize the complexity of the disease in ALS, but also reveal that UCHL1-eGFP reporter line would be a valuable tool to visualize and study various aspects of sensory nervous system development and degeneration in the context of numerous diseases.

No MeSH data available.


Related in: MedlinePlus

The epidermal innervation density is reduced only during disease end-stage in ALS.(A) UCHL1-eGFP reporter mice were crossed with hSOD1G93A mice to generate the hSOD1G93A-UeGFP ALS reporter mice and WT-UeGFP control mice. (B) eGFP+ axons are comparable at P30 (presymptomatic stage) between ALS reporter and control animals, but reveal a reduction in epidermal innervation density at P120 (end-stage) only in the hSOD1G93A-UeGFP mice. (C) Bar graph representation of average number of eGFP+ nerves per mm length of footpad epidermis in WT-UeGFP and hSOD1G93A-UeGFP mice at P30, P60, P90 and P120. (D) eGFP shows complete co-localization with UCHL1 expression in the hindlimb footpad of P120 (end-stage) hSOD1G93A-UeGFP mice. Bar graphs represent mean ± SEM. Student’s t-test, * P < 0.05. Scale bars B, D inset 50 μm; D 200 μm.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4519325&req=5

pone.0132815.g006: The epidermal innervation density is reduced only during disease end-stage in ALS.(A) UCHL1-eGFP reporter mice were crossed with hSOD1G93A mice to generate the hSOD1G93A-UeGFP ALS reporter mice and WT-UeGFP control mice. (B) eGFP+ axons are comparable at P30 (presymptomatic stage) between ALS reporter and control animals, but reveal a reduction in epidermal innervation density at P120 (end-stage) only in the hSOD1G93A-UeGFP mice. (C) Bar graph representation of average number of eGFP+ nerves per mm length of footpad epidermis in WT-UeGFP and hSOD1G93A-UeGFP mice at P30, P60, P90 and P120. (D) eGFP shows complete co-localization with UCHL1 expression in the hindlimb footpad of P120 (end-stage) hSOD1G93A-UeGFP mice. Bar graphs represent mean ± SEM. Student’s t-test, * P < 0.05. Scale bars B, D inset 50 μm; D 200 μm.

Mentions: By crossing UCHL1-eGFP female reporter line to male hSOD1G93A mice, we generated hSOD1G93A-UeGFP ALS reporter line [18] (Fig 6A). Peripheral neuropathy has been reported in some patients [31–36] and in ALS models [37,38]. To investigate the potential involvement of the sensory nervous system defects to disease pathology, and to determine the timing and extent of changes with respect to disease progression, we compared the innervation density of hind limb footpad in hSOD1G93A-UeGFP and WT-UeGFP mice, at P30 (pre-symptomatic), P60 (early-symptomatic), P90 (late-symptomatic) and P120 (end-stage) [39]. At P30, P60, and P90 the average number of intraepidermal nerve fibers innervating the skin were comparable between WT-UeGFP and hSOD1G93A-UeGFP mice (P30: 16% ± 1 axons in WT-UeGFP, n = 6 mice, 14 ± 1 axons in hSOD1G93A-UeGFP, n = 7 mice; P60: 21 ± 3 axons in WT-UeGFP, n = 3 mice, 16 ± 3 axons in hSOD1G93A-UeGFP, n = 3 mice; P90: 18 ± 2 axons in WT-UeGFP, n = 3 mice, 16 ± 2 axons in hSOD1G93A-UeGFP, n = 3 mice). However, by P120, skin innervation was significantly impaired only in the hSOD1G93A-UeGFP mice (Fig 6B and 6C; 18 ± 2 axons in WT-UeGFP, n = 6 mice; 12 ± 1 axons hSOD1G93A-UeGFP, n = 6 mice; P = 0.0254). The reduction in epidermal innervation density cannot be explained by reduced eGFP expression, because eGFP and UCHL1 are both present in the distal axons innervating the footpad (Fig 6D).


Visualization of Sensory Neurons and Their Projections in an Upper Motor Neuron Reporter Line.

Genç B, Lagrimas AK, Kuru P, Hess R, Tu MW, Menichella DM, Miller RJ, Paller AS, Özdinler PH - PLoS ONE (2015)

The epidermal innervation density is reduced only during disease end-stage in ALS.(A) UCHL1-eGFP reporter mice were crossed with hSOD1G93A mice to generate the hSOD1G93A-UeGFP ALS reporter mice and WT-UeGFP control mice. (B) eGFP+ axons are comparable at P30 (presymptomatic stage) between ALS reporter and control animals, but reveal a reduction in epidermal innervation density at P120 (end-stage) only in the hSOD1G93A-UeGFP mice. (C) Bar graph representation of average number of eGFP+ nerves per mm length of footpad epidermis in WT-UeGFP and hSOD1G93A-UeGFP mice at P30, P60, P90 and P120. (D) eGFP shows complete co-localization with UCHL1 expression in the hindlimb footpad of P120 (end-stage) hSOD1G93A-UeGFP mice. Bar graphs represent mean ± SEM. Student’s t-test, * P < 0.05. Scale bars B, D inset 50 μm; D 200 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519325&req=5

pone.0132815.g006: The epidermal innervation density is reduced only during disease end-stage in ALS.(A) UCHL1-eGFP reporter mice were crossed with hSOD1G93A mice to generate the hSOD1G93A-UeGFP ALS reporter mice and WT-UeGFP control mice. (B) eGFP+ axons are comparable at P30 (presymptomatic stage) between ALS reporter and control animals, but reveal a reduction in epidermal innervation density at P120 (end-stage) only in the hSOD1G93A-UeGFP mice. (C) Bar graph representation of average number of eGFP+ nerves per mm length of footpad epidermis in WT-UeGFP and hSOD1G93A-UeGFP mice at P30, P60, P90 and P120. (D) eGFP shows complete co-localization with UCHL1 expression in the hindlimb footpad of P120 (end-stage) hSOD1G93A-UeGFP mice. Bar graphs represent mean ± SEM. Student’s t-test, * P < 0.05. Scale bars B, D inset 50 μm; D 200 μm.
Mentions: By crossing UCHL1-eGFP female reporter line to male hSOD1G93A mice, we generated hSOD1G93A-UeGFP ALS reporter line [18] (Fig 6A). Peripheral neuropathy has been reported in some patients [31–36] and in ALS models [37,38]. To investigate the potential involvement of the sensory nervous system defects to disease pathology, and to determine the timing and extent of changes with respect to disease progression, we compared the innervation density of hind limb footpad in hSOD1G93A-UeGFP and WT-UeGFP mice, at P30 (pre-symptomatic), P60 (early-symptomatic), P90 (late-symptomatic) and P120 (end-stage) [39]. At P30, P60, and P90 the average number of intraepidermal nerve fibers innervating the skin were comparable between WT-UeGFP and hSOD1G93A-UeGFP mice (P30: 16% ± 1 axons in WT-UeGFP, n = 6 mice, 14 ± 1 axons in hSOD1G93A-UeGFP, n = 7 mice; P60: 21 ± 3 axons in WT-UeGFP, n = 3 mice, 16 ± 3 axons in hSOD1G93A-UeGFP, n = 3 mice; P90: 18 ± 2 axons in WT-UeGFP, n = 3 mice, 16 ± 2 axons in hSOD1G93A-UeGFP, n = 3 mice). However, by P120, skin innervation was significantly impaired only in the hSOD1G93A-UeGFP mice (Fig 6B and 6C; 18 ± 2 axons in WT-UeGFP, n = 6 mice; 12 ± 1 axons hSOD1G93A-UeGFP, n = 6 mice; P = 0.0254). The reduction in epidermal innervation density cannot be explained by reduced eGFP expression, because eGFP and UCHL1 are both present in the distal axons innervating the footpad (Fig 6D).

Bottom Line: Our findings corroborate previous studies, and suggest that more than five months is required for significant skin denervation.Crossing UCHL1-eGFP with hSOD1G93A mice generated hSOD1G93A-UeGFP reporter line of amyotrophic lateral sclerosis, and revealed sensory nervous system defects, especially towards disease end-stage.Our studies not only emphasize the complexity of the disease in ALS, but also reveal that UCHL1-eGFP reporter line would be a valuable tool to visualize and study various aspects of sensory nervous system development and degeneration in the context of numerous diseases.

View Article: PubMed Central - PubMed

Affiliation: Davee Department of Neurology and Clinical Neurological Sciences, Northwestern University, Feinberg School of Medicine, Chicago, IL, United States of America.

ABSTRACT
Visualization of peripheral nervous system axons and cell bodies is important to understand their development, target recognition, and integration into complex circuitries. Numerous studies have used protein gene product (PGP) 9.5 [a.k.a. ubiquitin carboxy-terminal hydrolase L1 (UCHL1)] expression as a marker to label sensory neurons and their axons. Enhanced green fluorescent protein (eGFP) expression, under the control of UCHL1 promoter, is stable and long lasting in the UCHL1-eGFP reporter line. In addition to the genetic labeling of corticospinal motor neurons in the motor cortex and degeneration-resistant spinal motor neurons in the spinal cord, here we report that neurons of the peripheral nervous system are also fluorescently labeled in the UCHL1-eGFP reporter line. eGFP expression is turned on at embryonic ages and lasts through adulthood, allowing detailed studies of cell bodies, axons and target innervation patterns of all sensory neurons in vivo. In addition, visualization of both the sensory and the motor neurons in the same animal offers many advantages. In this report, we used UCHL1-eGFP reporter line in two different disease paradigms: diabetes and motor neuron disease. eGFP expression in sensory axons helped determine changes in epidermal nerve fiber density in a high-fat diet induced diabetes model. Our findings corroborate previous studies, and suggest that more than five months is required for significant skin denervation. Crossing UCHL1-eGFP with hSOD1G93A mice generated hSOD1G93A-UeGFP reporter line of amyotrophic lateral sclerosis, and revealed sensory nervous system defects, especially towards disease end-stage. Our studies not only emphasize the complexity of the disease in ALS, but also reveal that UCHL1-eGFP reporter line would be a valuable tool to visualize and study various aspects of sensory nervous system development and degeneration in the context of numerous diseases.

No MeSH data available.


Related in: MedlinePlus