Neuronal STING activation is implicated in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). This study demonstrates that the STING pathway is activated in vulnerable cortical and spinal motor neurons in both familial and sporadic ALS, but not in less affected neurons. In a mouse model of C9orf72 repeat-associated ALS and FTD, STING activation is observed in layer V cortical motor neurons. The study also shows that STING activation occurs in a neuron-autonomous manner, with DNA damage serving as a key driver. Human iPSC-derived neurons harboring familial ALS mutations exhibit increased STING signaling, and elevated inflammatory markers can be suppressed with a STING inhibitor. The results suggest that STING activation in neurons contributes to the immunophenotype of ALS/FTD, characterized by innate immune signaling. The study further confirms that STING activation is specific to vulnerable neurons in ALS/FTD, as evidenced by immunohistochemical and immunofluorescence analyses. Additionally, STING signaling is increased in a C9orf72 mouse model, with activation occurring selectively in vulnerable deep layer V cortical motor neurons. The STING pathway is present and functional in primary mouse cortical neurons and healthy control human iPSC-derived neurons. SMNs and NGN2 neurons derived from a range of human fALS iPSCs show neuron-autonomous STING signaling activation. DNA damage induction through etoposide or glutamate treatment leads to γH2AX and STING pathway activation in primary mouse cortical neurons. TDP-43 depletion and C9orf72 DPR treatment each elicit DNA damage and STING pathway activation in iPSC-derived neurons. These findings highlight the role of STING activation in neuronal DNA damage and neuroinflammation in ALS and FTD.Neuronal STING activation is implicated in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). This study demonstrates that the STING pathway is activated in vulnerable cortical and spinal motor neurons in both familial and sporadic ALS, but not in less affected neurons. In a mouse model of C9orf72 repeat-associated ALS and FTD, STING activation is observed in layer V cortical motor neurons. The study also shows that STING activation occurs in a neuron-autonomous manner, with DNA damage serving as a key driver. Human iPSC-derived neurons harboring familial ALS mutations exhibit increased STING signaling, and elevated inflammatory markers can be suppressed with a STING inhibitor. The results suggest that STING activation in neurons contributes to the immunophenotype of ALS/FTD, characterized by innate immune signaling. The study further confirms that STING activation is specific to vulnerable neurons in ALS/FTD, as evidenced by immunohistochemical and immunofluorescence analyses. Additionally, STING signaling is increased in a C9orf72 mouse model, with activation occurring selectively in vulnerable deep layer V cortical motor neurons. The STING pathway is present and functional in primary mouse cortical neurons and healthy control human iPSC-derived neurons. SMNs and NGN2 neurons derived from a range of human fALS iPSCs show neuron-autonomous STING signaling activation. DNA damage induction through etoposide or glutamate treatment leads to γH2AX and STING pathway activation in primary mouse cortical neurons. TDP-43 depletion and C9orf72 DPR treatment each elicit DNA damage and STING pathway activation in iPSC-derived neurons. These findings highlight the role of STING activation in neuronal DNA damage and neuroinflammation in ALS and FTD.