Not likely. A directory is not a physical location on the disk. New contents for the directory are placed on disk based on their size and the availability of free space on the disk. The actual file for the directory is just a list of those locations.
However, it is very possible to have too many items in a directory. Once you get above a few hundred or thousand items (depending on the OS and type of access), access to the directory can cause a huge slowdown. If your directory has more than a few hundred items consider adding sub-folders to minimize the number in any given folder.
One final thought is that read errors could cause a large slow down if they are not so bad as to cause total failure. If you suspect this, run a diagnostic on the drive and get your data off ASAP if confirmed.
[Edit]
You mentioned above that these are radiology images. I assume with some form of patient ID or serial number/datestamp as the filename. Suggest you create folders for the first 2-3 letters of whatever denotes the ID and move the items into those. You can do this multiple folders deep and it will make finding the items no harder than it is now and greatly speed up directory indexing.
oldfolder
123456
123457
123558
...
55123
55124
...
99345
99346
newfolder
12
34
123456
123457
35
123558
...
55
12
55123
55124
...
99
34
99345
99346