Even though Route 38 maintains a pure east-west alignment as a whole, there are no fewer than four separate paths that Trainers can take between Ecruteak and Route 39. The northern-most path leads Trainers into the path of three other Trainers, two of which offer to give out their phone number for rematches at a later date. In Pokémon Crystal, one of these Trainers, Lass Dana, will also give Trainers who register her number an occasional Thunder Stone. Over half of this path is made up of grass patches.
The upper middle path is a branch of the northern path, separating near Dana and merging with the northern path near Route 39. There are no Trainers along this short stretch; there is, however, a patch of grass that is home to several species of wild Pokémon.
Unlike the two upper paths, which allow for two-way traffic, the lower paths are configured to allow only one direction of passage. The lower middle path is designed for travelers heading west to Olivine City and is a viable alternative to the grass-lined northern path. The path heads due west from the gate and travels down a fence-lined walkway that is home to one Trainer. Past this Trainer, there is a BerryGSC or ApricornHGSS bush just before a ledge that overlooks a grass-lined pathway. After hopping this ledge, Trainers are forced to head north along this passageway, encountering another Trainer before merging into the upper middle path.
The southern-most path is an "express route" from Olivine to Ecruteak and is designed so that only eastward movements are permitted. After breaking from the northern path near Route 39, the path heads south, passing a signpost before traversing a ledge that borders a T-intersection. At this junction, Trainers can either head north into the field of grass along the lower middle path or continue east along the southern edge of the ledge of the lower middle path and, eventually, along the fenced area to Ecruteak.
Trainers along this route may sometimes use Moomoo Milk in battles, in reference to the nearby Moomoo Farm.
Route 38 first appeared in Indubitably Ditto, where the Team Rocket Elite Trio tried to capture Suicune there, but they were put to sleep by Crystal's Parasee's Spore before they could reach it. When they woke up, Suicune was gone and Eusine appeared before them, challenging them to a battle. After defeating the trio, Eusine took the Clear Bell they had and headed for the Tin Tower, where Crystal was currently battling Suicune.
In Pleased as Punch With Parasect, Crystal and Silver were traversing through the route on their way to Ecruteak City. However, along the way, they were attacked by Team Rocket Executive Ariana, who was tasked with getting rid of any obstacles in Team Rocket's way. Using his Rhyperior and the Earth Plate, Silver managed to escape to Ecruteak to ask help from Morty in locating the remaining Plates.
In With a Little Help From Hitmonchan, Crystal was defeated by Ariana, shortly after which the other Executives, Archer, Petrel, and Proton, also showed up. Following Archer's prediction, Arceus also showed up on the route, having descended back to this world from its heavenly residence. Realizing that Arceus was a wild Pokémon, Crystal attempted to capture it. After a Level Ball had failed to have any effect on the Alpha Pokémon, Crystal used a Heavy Ball, which surprisingly managed to contain Arceus for a few seconds before it escaped. When Arceus fled, Crystal gave pursuit, but was soon blasted down from the sky by Arceus.
Pokémon Gold & Silver: The Golden Boys
Route 38 appeared in A Promise Given to Miltank. After Whitney's Miltank had been diagnosed with a deadly disease, Gold, Whitney, and Morty set out to find the Berries required to cure her, only having time until sunset before the disease would claim Miltank's life. Their Berry-gathering was hampered by wild Pokémon appearing from the Berry trees, but ultimately, they managed to find a tree with lots of Berries on it. Gold used his Sudowoodo to lure away the Heracross that were swarming around the tree, while Whitney had Pikachu knock the Berries down with Headbutt. The Berries were delivered to the Ecruteak City Pokémon Center just in time, saving Miltank's life. Out of gratefulness, Whitney gave Gold a Plain Badge and returned his Pikachu to him.